Substituted bridged urea analogs as sirtuin modulators

ABSTRACT

The present invention relates to novel substituted bridged urea analog compounds of Formula (I) or pharmaceutically acceptable salts thereof, corresponding pharmaceutical compositions, processes for making and use of such compounds, alone or in combination with other therapeutic agents, as Sirtuin Modulators useful for increasing lifespan of a cell, and in treating and/or preventing a wide variety of diseases and disorders, which include, but are not limited to, for example, diseases or disorders related to aging or stress, diabetes, obesity, neurodegenerative diseases, cardiovascular disease, blood clotting disorders, inflammation, cancer, and/or flushing as well as diseases or disorders that would benefit from increased mitochondrial activity.

FIELD OF THE INVENTION

In general, the present invention relates to substituted bridged ureaanalog compounds of Formulas (I) to (VI), corresponding anlogs orderivatives thereof, or pharmaceutically acceptable salts thereof,corresponding pharmaceutical compositions, processes for making andmethods or uses of such compounds, alone or in combination with othertherapeutic agents, as Sirtuin Modulators useful for increasing lifespanof a cell, and in treating and/or preventing a wide variety of diseasesand disorders, which include, but are not limited to, for example,diseases or disorders related to aging or stress, diabetes, obesity,neurodegenerative diseases, cardiovascular disease, blood clottingdisorders, inflammation, cancer, and/or flushing as well as diseases ordisorders that would benefit from increased mitochondrial activity.

BACKGROUND

The Silent Information Regulator (SIR) family of genes represents ahighly conserved group of genes present in the genomes of organismsranging from archaebacteria to eukaryotes. The encoded SIR proteins areinvolved in diverse processes from regulation of gene silencing to DNArepair. A well-characterized gene in this family is S. cerevisiae SIR2,which is involved in silencing HM loci that contain informationspecifying yeast mating type, telomere position effects and cell aging.The yeast Sir2 protein belongs to a family of histone deacetylases. Theproteins encoded by members of the SIR gene family show high sequenceconservation in a 250 amino acid core domain. The Sir2 homolog, CobB, inSalmonella typhimurium, functions as an NAD (nicotinamide adeninedinucleotide)-dependent ADP-ribosyl transferase.

The Sir2 protein is a class III deacetylase which uses NAD as acosubstrate. Unlike other deacetylases, many of which are involved ingene silencing, Sir2 is insensitive to class I and II histonedeacetylase inhibitors like trichostatin A (TSA).

Deacetylation of acetyl-lysine by Sir2 is tightly coupled to NADhydrolysis, producing nicotinamide and a novel acetyl-ADP ribosecompound. The NAD-dependent deacetylase activity of Sir2 is essentialfor its functions, which can connect its biological role with cellularmetabolism in yeast. Mammalian Sir2 homologs have NAD-dependent histonedeacetylase activity.

Biochemical studies have shown that Sir2 can readily deacetylate theamino-terminal tails of histones H3 and H4, resulting in the formationof 2′/3′-O-acetyl-ADP-ribose (OAADPR) and nicotinamide. Strains withadditional copies of SIR2 display increased rDNA silencing and a 30%longer life span. It has also been shown that additional copies of theC. elegans SIR2 homolog, sir-2.1, and the D. melanogaster dSir2 geneextend life span in those organisms. This implies that theSIR2-dependent regulatory pathway for aging arose early in evolution andhas been well conserved. Today, Sir2 genes are believed to have evolvedto enhance an organism's health and stress resistance to increase itschance of surviving adversity.

In humans, there are seven Sir2-like genes (SIRT1-SIRT7) that share theconserved catalytic domain of Sir2. SIRT1 is a nuclear protein with thehighest degree of sequence similarity to Sir2. SIRT1 regulates multiplecellular targets by deacetylation including the tumor suppressor p53,the cellular signaling factor NF-κB, and the FOXO transcription factor.

SIRT3 is a homolog of SIRT1 that is conserved in prokaryotes andeukaryotes. The SIRT3 protein is targeted to the mitochondrial cristaeby a unique domain located at the N-terminus. SIRT3 has NAD⁺-dependentprotein deacetylase activity and is ubiquitously expressed, particularlyin metabolically active tissues. Upon transfer to the mitochondria,SIRT3 is believed to be cleaved into a smaller, active form by amitochondrial matrix processing peptidase (MPP).

Caloric restriction has been known for over 70 years to improve thehealth and extend the lifespan of mammals. Yeast life span, like that ofmetazoans, is also extended by interventions that resemble caloricrestriction, such as low glucose. The discovery that both yeast andflies lacking the SIR2 gene do not live longer when caloricallyrestricted provides evidence that SIR2 genes mediate the beneficialhealth effects of a restricted calorie diet. Moreover, mutations thatreduce the activity of the yeast glucose-responsive cAMP (adenosine3′,5′-monophosphate)-dependent (PKA) pathway extend life span in wildtype cells but not in mutant sir2 strains, demonstrating that SIR2 islikely to be a key downstream component of the caloric restrictionpathway.

In addition to therapeutic potential, structural and biophysical studiesof SIRT1 activity and activation by small molecule sirtuin modualtorswould be useful to advance understanding of the biological function ofsirtuins, to further the understanding of the mechanism of action ofsirtuin activation and to aid in the development of assays that identifynovel sirtuin modulators.

The present invention is directed to overcoming these and other problemsencountered in the art.

SUMMARY OF THE INVENTION

In general, the present invention relates to substituted bridged ureaanalog compounds of Formulas (I) to (VI), corresponding anlogs orderivatives thereof, or pharmaceutically acceptable salts thereof,corresponding pharmaceutical compositions, processes for making and useof such compounds, alone or in combination with other therapeuticagents, as Sirtuin Modulators useful for increasing lifespan of a cell,and in treating and/or preventing a wide variety of diseases anddisorders, which include, but are not limited to, for example, diseasesor disorders related to aging or stress, diabetes, obesity,neurodegenerative diseases, cardiovascular disease, blood clottingdisorders, inflammation, cancer, and/or flushing as well as diseases ordisorders that would benefit from increased mitochondrial activity.

In particular, the present invention relates to novel compounds ofFormulas (I) to (VI), corresponding analogs (i.e., with hydrogensubstitution at the R² position) and corresponding pharmaceuticalcompositions comprising compounds of Formulas (I) to (VI) respectively.

The present invention also relates to processes for making compounds ofFormulas (I) to (VI), and corresponding analogs (i.e., with hydrogensubstitution at the R² position), respectively.

The present invention also relates to methods for using or uses ofSirtuin Modulator compounds as defined herein in treating and/orpreventing a wide variety of diseases and disorders, which include, butare not limited to, for example, diseases or disorders related to agingor stress, diabetes, obesity, neurodegenerative diseases, cardiovasculardisease, blood clotting disorders, inflammation, cancer, and/or flushingas well as diseases or disorders that would benefit from increasedmitochondrial activity, further which may be selected from or include,but are not limited to psoriasis, atopic dermatitis, acne, rosacea,inflammatory bowel disease, osteoporosis, sepsis, arthritis, COPD,systemic lupus erythematosus and ophthalmic inflammation.

DETAILED DESCRIPTION OF THE INVENTION

In general, the present invention relates to substituted bridged ureaanalog compounds of Formulas (I) to (VI), corresponding anlogs orderivatives thereof, or pharmaceutically acceptable salts thereof,corresponding pharmaceutical compositions, processes for making and useof such compounds, alone or in combination with other therapeuticagents, as Sirtuin Modulators useful for increasing lifespan of a cell,and in treating and/or preventing a wide variety of diseases anddisorders, which include, but are not limited to, for example, diseasesor disorders related to aging or stress, diabetes, obesity,neurodegenerative diseases, cardiovascular disease, blood clottingdisorders, inflammation, cancer, and/or flushing as well as diseases ordisorders that would benefit from increased mitochondrial activity.

Compounds

In particular, the present invention relates to novel compounds ofFormulas (I) to (VI), corresponding analogs (i.e., with hydrogensubstitution at the R² position) and corresponding pharmaceuticalcompositions comprising compounds of Formulas (I) to (VI), respectively.

International Patent Application No. WO09/061879, International FilingDate: 13 May 2014 discloses novel sirtuin-modulating substituted bridgedurea and related analogs compounds of Formula (I):

ora pharmaceutically acceptable salt thereof, corresponding pharmaceuticalcompositions, combinations with other therapeutic agents, methods formaking and methods or uses for increasing the lifespan of a cell, andtreating and/or preventing a wide variety of diseases and disordersincluding, for example, diseases or disorders related to aging orstress, diabetes, obesity, neurodegenerative diseases, cardiovasculardisease, blood clotting disorders, inflammation, cancer, and/or flushingas well as diseases or disorders that would benefit from increasedmitochondrial activity.

In one aspect, the present invention provides novel sirtuin-modulatingcompounds of Structural Formulas (I) to (VI), respectively correspondinganalogs (i.e., with hydrogen substitution at the R² position) as aredescribed in detail below.

In one aspect, the present invention relates to a compound of Formula(I):

where:

X₁ or X₂ independently is selected from —N or —C;

R¹ is hydrogen, halogen, —CN, carbocyclyl, heterocyclyl, —N-substitutedheterocyclyl, aryl, heteroaryl, —C(O)R_(a) or —C(O)—NR_(b)R_(c);

R² is halogen, -straight or branched C₁-C₆ alkyl, -straight orbranched-C₁-C₆ haloalkyl, or —C(O)—NR_(b)R_(c);

R³ is hydrogen, halogen, -hydroxy, -straight or branched C₁-C₆ alkyl, or-straight or branched-C₁-C₆ haloalkyl;

R⁴ is hydrogen or —C(O)NR_(b)R_(c);

where:

-   -   when X₂ is —N, R₂ is non-existent; or    -   when X₂ is —C, R₂ is as defined above;    -   each R¹, R², R³ or R⁴ as defined above optionally is further        substituted with one or more substituents selected from        hydrogen, halogen, —OH, —(CH₂)_(x)OH, —C≡N, —NR_(d)R_(e),        -straight or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆        haloalkyl, -straight or branched C₁-C₆ alkoxy, -straight or        branched C₁-C₆ haloalkoxy, —O-straight or branched-C₁-C₆        haloalkyl, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,        heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,        —O—(CH₂)_(x)CH(OH)CH₂(OH), or —C(O)OR_(f);    -   each R_(a), R_(b), R_(c), R_(d), R_(e), or R_(f) as defined        above independently is selected from hydrogen, -straight or        branched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,        —C₁-C₆-cycloalkyl, —(CH₂)_(x)C₁-C₆-cycloalkyl, heterocyclyl, —N—        heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl,        —(CHR_(g))_(x)heteroaryl;    -   where:        -   R_(g) is -straight or branched C₁-C₆ alkyl, -straight or            branched-C₁-C₆ haloalkyl;        -   each R_(a), R_(b), R_(c) R_(d), R_(e), or R_(f) as defined            above optionally is further substituted with one or more            substituents selected from hydrogen, halogen, —OH, —C≡N,            -straight or branched C₁-C₆ alkyl, -straight or            branched-C₁-C₆ haloalkyl, -straight or branched C₁-C₆            alkoxy, —O-straight or branched-C_(r) C₆ haloalkyl, —C₁-C₆            cycloalkyl, carbocyclyl, —(CH₂)_(x)-carbocyclyl,            -heterocyclyl, —O-heterocyclyl aryl, -heteroaryl,            —(CH₂)_(x)-heteroaryl, —O—(CH₂)_(x)CH(OH)CH₂(OH),            —(CH₂)_(x)—OH, or —C(O)—OH;

m is an integer from 1 to 3;

n is an integer selected from 1 to 3;

x is 0 or an integer from 1 to 6; or

a pharmaceutically salt thereof.

In another aspect, the present invention relates to a compound of thepresent invention as defined above (i.e., compounds of StructuralFormulas (I) to (VI), respectively corresponding analogs (i.e., withhydrogen substitution at the R² position) and throughout the instantapplication, where it is provided that:

-   -   when n=1, m≠1; and    -   when n=3, m≠3.

In another aspect, the present invention relates to a compound of thepresent invention, where R² is C(O)—NR_(b)R_(c); wherein R_(b) and R_(e)are as defined above and throughout the present application.

In another aspect, the present invention relates to a compound ofFormulas (I) to (VI), where:

m is 1;

n is 2 or 3; and

R⁴ is hydrogen.

In another aspect, the present invention relates to a compound ofFormulas (I) to (VI), where:

m is 1;

n is 2 or 3; and

R⁴ is —C(O)NR_(b)R_(c), wherein each R_(b) and R_(e) is as definedabove.

In another aspect, the present invention relates to a compound ofFormulas (I) to (VI), where:

m is 1;

n is 3; and

R⁴ is R⁴ is hydrogen or —C(O)NR_(b)R_(c), wherein R_(b) and R_(e) is asdefined above in claim 1.

In one aspect, the present invention relates to a compound of Formula(II):

where:

X₁ or X₂ independently is selected from —N or —C;

R¹ is hydrogen, halogen, —CN, carbocyclyl, heterocyclyl, —N-substitutedheterocyclyl, aryl or heteroaryl.

R² is halogen, -straight or branched C₁-C₆ alkyl, -straight orbranched-C₁-C₆ haloalkyl, or —C(O)—NR_(b)R_(c);

R³ is hydrogen, halogen, -hydroxy, -straight or branched C₁-C₆ alkyl, or-straight or branched-C₁-C₆ haloalkyl;

R⁴ is hydrogen or —C(O)NR_(b)R_(c);

where:

-   -   when X₂ is —N, R₂ is non-existent; or    -   when X₂ is —C, R₂ is as defined above;    -   each R¹, R², R³ or R⁴ as defined above optionally is further        substituted with one or more substituents selected from        hydrogen, halogen, —OH, —(CH₂)_(x)OH, —C≡N, —NR_(d)R_(e),        -straight or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆        haloalkyl, -straight or branched C₁-C₆ alkoxy, -straight or        branched C₁-C₆ haloalkoxy, —O-straight or branched-C₁-C₆        haloalkyl, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,        heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,        —O—(CH₂)_(x)CH(OH)CH₂(OH), or —C(O)OR_(f);    -   each R_(a), R_(b), R_(c), R_(d), R_(e), or R_(f) as defined        above independently is selected from hydrogen, -straight or        branched C₁-C₆ alkyl, -straight or branched-C₁-C₆haloalkyl,        —C₁-C₆-cycl ° alkyl, —(CH₂)—C₁-C₆-cycl ° alkyl, heterocyclyl,        —N— heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl,        —(CHR_(g))_(x)heteroaryl;    -   where:        -   R_(g) is -straight or branched C₁-C₆ alkyl, -straight or            branched-C₁-C₆ haloalkyl;        -   each R_(a), R_(b), R_(c) R_(d), R_(e), or R_(f) as defined            above optionally is further substituted with one or more            substituents selected from hydrogen, halogen, —OH, —C≡N,            -straight or branched C₁-C₆ alkyl, -straight or            branched-C₁-C₆ haloalkyl, -straight or branched C₁-C₆            alkoxy, —O-straight or branched-C₁-C₆ haloalkyl, —C₁-C₆            cycloalkyl, carbocyclyl, —(CH₂)_(x)-carbocyclyl,            -heterocyclyl, —O-heterocyclyl aryl, -heteroaryl,            —(CH₂)_(x)-heteroaryl, —O—(CH₂)_(x)CH(OH)CH₂(OH),            —(CH₂)_(x)—OH, or —C(O)—OH;

m is an integer from 1 to 3;

n is an integer selected from 1 to 3;

x is 0 or an integer from 1 to 6; or

a pharmaceutically salt thereof.

In one aspect, the present invention relates to a compound of Formula(III):

where:

X₁ or X₂ independently is selected from —N or —C;

R¹ is —C(O)R_(a) or —C(O)—NR_(b)R_(c);

R² is halogen, -straight or branched C₁-C₆ alkyl, -straight orbranched-C₁-C₆ haloalkyl, or —C(O)—NR_(b)R_(c);

R³ is hydrogen, halogen, -hydroxy, -straight or branched C₁-C₆ alkyl, or-straight or branched-C₁-C₆ haloalkyl;

R⁴ is hydrogen or —C(O)NR_(b)R_(c);

where:

-   -   when X₂ is —N, R₂ is non-existent; or    -   when X₂ is —C, R₂ is as defined above;    -   each R¹, R², R³ or R⁴ as defined above optionally is further        substituted with one or more substituents selected from        hydrogen, halogen, —OH, —(CH₂)_(x)OH, —C≡N, —NR_(d)R_(e),        -straight or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆        haloalkyl, -straight or branched C₁-C₆ alkoxy, -straight or        branched C₁-C₆ haloalkoxy, —O-straight or branched-C₁-C₆        haloalkyl, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,        heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,        —O—(CH₂)_(x)CH(OH)CH₂(OH), or —C(O)OR_(f);    -   each R_(a), R_(b), R_(c), R_(d), R_(e), or R_(f) as defined        above independently is selected from hydrogen, -straight or        branched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,        —C₁-C₆-cycloalkyl, —(CH₂)_(x)C₁-C₆-cycloalkyl, heterocyclyl, —N—        heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl,        —(CHR_(g))_(x)heteroaryl;    -   where:        -   R_(g) is -straight or branched C₁-C₆ alkyl, -straight or            branched-C₁-C₆ haloalkyl;        -   each R_(a), R_(b), R_(c) R_(d), R_(e), or R_(f) as defined            above optionally is further substituted with one or more            substituents selected from hydrogen, halogen, —OH, —C≡N,            -straight or branched C₁-C₆ alkyl, -straight or            branched-C₁-C₆ haloalkyl, -straight or branched C₁-C₆            alkoxy, —O-straight or branched-C₁-C₆ haloalkyl, —C₁-C₆            cycloalkyl, carbocyclyl, —(CH₂)_(x)-carbocyclyl,            -heterocyclyl, —O-heterocyclyl aryl, -heteroaryl,            —(CH₂)_(x)-heteroaryl, —O—(CH₂)_(x)CH(OH)CH₂(OH),            —(CH₂)_(x)—OH, or —C(O)—OH;

m is an integer from 1 to 3;

n is an integer selected from 1 to 3;

x is 0 or an integer from 1 to 6; or

a pharmaceutically salt thereof.

In another aspect, the present invention relates to a compound of thepresent invention as defined above (i.e., compounds of StructuralFormulas (I) to (VI), respectively corresponding analogs (i.e., withhydrogen substitution at the R² position) and throughout the instantapplication, where it is provided that:

-   -   when n=1, m≠1; and    -   when n=3, m≠3.

In another aspect, the present invention relates to a compound of thepresent invention as described herein, where R² is C(O)—NR_(b)R_(c);wherein R_(b) and R_(c) are as defined above and throughout the presentapplication.

In another aspect, the present invention relates to a compound ofFormulas (I) to (VI) or any compound as described herein, where:

m is 1;

n is 2 or 3; and

R⁴ is hydrogen.

In another aspect, the present invention relates to a compound ofFormulas (I) to (VI), where:

m is 1;

n is 2 or 3; and

R⁴ is —C(O)NR_(b)R_(c), wherein each R_(b) and R_(c) is as definedabove.

In another aspect, the present invention relates to a compound ofFormula (IV):

where:

X₁ or X₂ independently is selected from —N or —C;

where:

-   -   when X₂ is —N, R₂ is non-existent; or    -   when X₂ is —C, R₂ is as defined above;

R¹ is hydrogen, halogen, —CN, carbocyclyl, heterocyclyl, —N-substitutedheterocyclyl, aryl, heteroaryl, —C(O)R_(a) or —C(O)—NR_(b)R_(c);

R² is halogen, -straight or branched C₁-C₆ alkyl, -straight orbranched-C₁-C₆ haloalkyl, or —C(O)—NR_(b)R_(c);

R³ is hydrogen, halogen, -hydroxy, -straight or branched C₁-C₆ alkyl, or-straight or branched-C₁-C₆ haloalkyl;

each R⁵ and R⁶ independently is selected from hydrogen, -straight orbranched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,—C₁-C₆cycloalkyl, —(CH₂)_(x)C₁-C₆cycloalkyl, heterocyclyl,—N-heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl,—(CHR_(g))_(x)heteroaryl;

wherein:

-   -   each R¹, R², R³, R⁵ and R⁶ as defined above optionally is        further substituted with one or more substituents selected from        hydrogen, halogen, —OH, —(CH₂)_(x)OH, —C≡N, —NR_(d)R_(e),        -straight or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆        haloalkyl, -straight or branched C₁-C₆ alkoxy, -straight or        branched C₁-C₆ haloalkoxy, —O-straight or branched-C₁-C₆        haloalkyl, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,        heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,        —O—(CH₂)_(x)CH(OH)CH₂(OH), or —C(O)OR_(f);    -   each R_(a), R_(b), R_(c), R_(d), R_(e), R_(f) or R_(g) as        defined above independently is selected from hydrogen, -straight        or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,        —C₁-C₆-cycloalkyl, —(CH₂)_(x)C₁-C₆-cycloalkyl, heterocyclyl, —N—        heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl;    -   where:        -   each R_(a), R_(b), R_(c) R_(d), R_(e), R_(f) or R_(g) as            defined above optionally is further substituted with one or            more substituents selected from hydrogen, halogen, —OH,            —(CH₂)_(x)OH, —C≡N, NR_(h)R_(i), -straight or branched C₁-C₆            alkyl, -straight or branched-C₁-C₆ haloalkyl, -straight or            branched C₁-C₆ alkoxy, -straight or branched-C₁-C₆            haloalkoxy, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,            heterocyclyl, -heterocyclyl, —O-heterocyclyl, aryl,            -heteroaryl, —(CH₂)_(x)-heteroaryl,            —O—(CH₂)_(x)CH(OH)CH₂(OH), —(CH₂)_(x)—OH, or —C(O)OR_(j);        -   where:        -   each Rh, Ri and Rj independently is selected from hydrogen,            -straight or branched C₁-C₆ alkyl or -straight or            branched-C₁-C₆ haloalkyl;

m is an integer from 1 to 3;

n is an integer selected from 2 to 3;

x is 0 or an integer from 1 to 6; or

a pharmaceutically salt thereof.

In another aspect, the present invention relates to a compound of thepresent invention, where n is 2 or 3 and m is 1.

In another aspect, the present invention relates to a compound ofFormula (V):

where:

R¹ is hydrogen, halogen, —CN, carbocyclyl, heterocyclyl, —N-substitutedheterocyclyl, aryl, heteroaryl, —C(O)R_(a) or —C(O)—NR_(b)R_(c);

R² halogen, -straight or branched C₁-C₆ alkyl, -straight orbranched-C₁-C₆ haloalkyl, or —C(O)—NR_(b)R_(c);

R³ is hydrogen, halogen, -hydroxy, -straight or branched C₁-C₆ alkyl, or-straight or branched-C₁-C₆ haloalkyl;

R4 is hydrogen or —C(O)—NR_(b)R_(c);

where:

-   -   each R¹, R², R³ or R⁴ as defined above optionally is further        substituted with one or more substituents selected from        hydrogen, halogen, —OH, —(CH₂)_(x)OH, —C≡N, —NR_(d)R_(e),        -straight or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆        haloalkyl, -straight or branched C₁-C₆ alkoxy, -straight or        branched C₁-C₆ haloalkoxy, —O-straight or branched-C₁-C₆        haloalkyl, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,        heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,        —O—(CH₂)_(x)CH(OH)CH₂(OH), or —C(O)OR_(f);    -   each R_(a), R_(b), R_(e), R_(d), R_(e), or R_(f) as defined        above independently is selected from hydrogen, -straight or        branched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,        —C₁-C₆-cycloalkyl, —(CH₂)_(x)C₁-C₆-cycloalkyl, heterocyclyl, —N—        heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl;    -   where:        -   each R_(a), R_(b), R_(c) R_(d), R_(e), or R_(f) as defined            above optionally is further substituted with one or more            substituents selected from hydrogen, halogen, —OH,            —(CH₂)_(x)OH, —C≡N, —NR_(g)R_(h), -straight or branched            C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,            -straight or branched C₁-C₆ alkoxy, -straight or            branched-C₁-C₆ haloalkoxy, —C₁-C₆ cycloalkyl,            —(CH₂)_(x)-cycloalkyl, heterocyclyl, -heterocyclyl,            —O-heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,            —O—(CH₂)_(x)CH(OH)CH₂(OH), —(CH₂)_(x)—OH, or —C(O)OR_(i);        -   wherein:        -   each R_(g), R_(h) and R_(i) as defined above independently            is selected from hydrogen, -straight or branched C₁-C₆ alkyl            or -straight or branched-C₁-C₆ haloalkyl;

m is an integer from 1 to 3;

n is an integer selected from 2 to 3;

x is 0 or an integer from 1 to 6; or

a pharmaceutically salt thereof;

In another aspect, the present invention relates to a compound ofFormula (IV):

where:

R¹ is hydrogen, halogen, —CN, carbocyclyl, heterocyclyl, —N-substitutedheterocyclyl, aryl, heteroaryl, —C(O)R_(a) or —C(O)—NR_(b)R_(c);

R² halogen, -straight or branched C₁-C₆ alkyl, -straight orbranched-C₁-C₆ haloalkyl, or —C(O)—NR_(b)R_(c);

R³ is hydrogen, halogen, -hydroxy, -straight or branched C₁-C₆ alkyl, or-straight or branched-C₁-C₆ haloalkyl;

each R⁵ and R⁶ independently is selected from hydrogen, -straight orbranched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,—C₁-C₆cycloalkyl, —(CH₂)_(x)C₁-C₆cycloalkyl, heterocyclyl,—N-heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl,—(CHR_(g))_(x)heteroaryl;

where:

-   -   each R¹, R², R³, R⁵ and R⁶ as defined above optionally is        further substituted with one or more substituents selected from        hydrogen, halogen, —OH, —(CH₂)_(x)OH, —C≡N, —NR_(d)R_(e),        -straight or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆        haloalkyl, -straight or branched C₁-C₆ alkoxy, -straight or        branched C₁-C₆ haloalkoxy, —O-straight or branched-C₁-C₆        haloalkyl, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,        heterocyclyl, aryl, -heteroaryl, —(CH₂)_(x)-heteroaryl,        —O—(CH₂)_(x)CH(OH)CH₂(OH), or —C(O)OR_(f);    -   each R_(a), R_(b), R_(c), R_(d), R_(e), R_(f) or R_(g) as        defined above independently is selected from hydrogen, -straight        or branched C₁-C₆ alkyl, -straight or branched-C₁-C₆ haloalkyl,        —C₁-C₆-cycloalkyl, —(CH₂)_(x)C₁-C₆-cycloalkyl, heterocyclyl, —N—        heterocyclyl, aryl, heteroaryl, or —(CH₂)_(x)heteroaryl;    -   where:        -   each R_(a), R_(b), R_(c) R_(d), R_(e), R_(f) or R_(g) as            defined above optionally is further substituted with one or            more substituents selected from hydrogen, halogen, —OH,            —(CH₂)_(x)OH, —C≡N, NR_(h)R_(i), -straight or branched C₁-C₆            alkyl, -straight or branched-C₁-C₆ haloalkyl, -straight or            branched C₁-C₆ alkoxy, -straight or branched-C₁-C₆            haloalkoxy, —C₁-C₆ cycloalkyl, —(CH₂)_(x)-cycloalkyl,            heterocyclyl, -heterocyclyl, —O-heterocyclyl, aryl,            -heteroaryl, —(CH₂)_(x)-heteroaryl,            —O—(CH₂)_(x)CH(OH)CH₂(OH), —(CH₂)_(x)—OH, or —C(O)OR_(j);        -   where:        -   each Rh, Ri and Rj independently is selected from hydrogen,            -straight or branched C₁-C₆ alkyl or -straight or            branched-C₁-C₆ haloalkyl;

m is an integer from 1 to 3;

n is an integer selected from 2 to 3;

x is 0 or an integer from 1 to 6; or

a pharmaceutically salt thereof;

In another aspect, the present invention relates to compounds ofFormulas (I) to (IV), respectively, wherein R¹ is selected from:

In another aspect, the present invention relates to compound(s) ofFormulas (I) to (IV), respectively, R1 is

In another aspect, the present invention relates to compound(s) ofFormulas (I) to (IV), respectively, where R⁴ is selected from:

In another aspect, the present invention relates to compound(s) ofFormulas (I) to (IV), respectively, R4 is

In another aspect, the present invention relates to a compound which isas defined in Table 1 of the instant specification starting at page 303:

In another aspect, the present invention relates to a compound which is:

CHART 3 Chemical Name: Generated by Structure ChemAxon

(9S)-N-(5-fluoropyridin-2-yl)-5- [(3S)-3-(trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene-4-carboxamide

(9S)-N-(pyrazin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-4-carboxamide

(9S)-N-(pyridin-3-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-4-carboxamide

(9S)-N-(pyridin-3-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-4-carboxamide

(9S)-N-(pyrazin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-4-carboxamide

In another aspect, the present invention relates to a compound which isa corresponding analog or derivative of the present invention s (i.e.,with hydrogen substitution at the R² position):

CHART 4 Three Carbon Unit and Pyridine-No Meta Substitution ChemicalName: Generated by Structure ChemAxon

(9S)-N-(pyridin-2-yl)-5-[(2S)-2- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(5-fluoropyridin-3-yl)-5- [2-(trifluoromethyl)pyridin-4-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrimidin-5-yl)-5-[5- (trifluoromethyl)pyridin-3-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-5-(2-methylpyridin-4-yl)-N- (pyridin-2-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-2-yl)-5-[(2R)-2- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(5-fluoropyridin-3-yl)-5- (2-methylpyridin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2,4,6-triene-8-carboxamide

(9S)-N-(pyridazin-3-yl)-5-[2- (trifluoromethyl)pyridin-4-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-2-yl)-5-[2- (trifluoromethyl)pyridin-4-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-2-yl)-5-[(3S)-3- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrimidin-4-yl)-5-[2- (trifluoromethyl)pyridin-4-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrimidin-5-yl)-5-[2- (trifluoromethyl)pyridin-4-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-5-(2-methylpyridin-4-yl)-N- (pyrimidin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-5-(2-methylpyridin-4-yl)-N- (pyrazin-2-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-5-(2-methylpyridin-4-yl)-N- (pyridazin-3-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(2-methyl-2H-indazol-5- yl)-5-(2-methylpyridin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2,4,6-triene-8-carboxamide

(9S)-5-(2-methylpyridin-4-yl)-N- (pyrimidin-5-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrazin-2-yl)-5-[2- (trifluoromethyl)pyridin-4-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrimidin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(dimethyl-1,3-thiazol-2- yl)-5-(2-methylpyridin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(6-methoxypyrazin-2-yl)- 5-(2-methylpyridin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(6-methylpyrazin-2-yl)-5- (2-methylpyridin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrimidin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca - 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(5-fluoropyridin-2-yl)-5- [(3S)-3-(trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-3-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(1-methyl-1H-pyrazol-4- yl)-5-(2-methylpyridin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrazin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-3-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(5-fluoropyridin-2-yl)-5- [(3S)-3-(trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene-8-carboxamide

(9S)-N-(pyrazin-2-yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamide

(9S)-N-(pyridin-2-yl)-5-[(3S)-3- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca- 2(7),3,5-triene-8-carboxamidehydrochloride

Terms and Definitions Section 1

Certain compounds of the present invention may exist in particulargeometric or stereoisomeric forms. The present invention contemplatesall such compounds, including cis- and trans-isomers, (R)- and(S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, the racemicmixtures thereof, and other mixtures thereof, as falling within thescope of the invention. Additional asymmetric carbon atoms may bepresent in a substituent such as an alkyl group. All such isomers, aswell as mixtures thereof, are intended to be included in this invention.

The compounds and salts thereof described herein can also be present asthe corresponding hydrates (e.g., hemihydrate, monohydrate, dihydrate,trihydrate, tetrahydrate) or solvates. Suitable solvents for preparationof solvates and hydrates can generally be selected by a skilled artisan.

The compounds and salts thereof can be present in amorphous orcrystalline (including co-crystalline and polymorph) forms.

Sirtuin-modulating compounds of the invention advantageously modulatethe level and/or activity of a sirtuin protein, particularly thedeacetylase activity of the sirtuin protein.

Separately or in addition to the above properties, certainsirtuin-modulating compounds of the invention do not substantially haveone or more of the following activities: inhibition of PI3-kinase,inhibition of aldoreductase, inhibition of tyrosine kinase,transactivation of EGFR tyrosine kinase, coronary dilation, orspasmolytic activity, at concentrations of the compound that areeffective for modulating the deacetylation activity of a sirtuin protein(e.g., such as a SIRT1 and/or a SIRT3 protein).

An “alkyl” group or “alkane” is a straight chained or branchednon-aromatic hydrocarbon which is completely saturated. Typically, astraight chained or branched alkyl group has from 1 to about 20 carbonatoms, preferably from 1 to about 10 unless otherwise defined. Examplesof straight chained and branched alkyl groups include methyl, ethyl,n-propyl, iso-propyl, n-butyl, sec-butyl, tert-butyl, pentyl, hexyl,pentyl and octyl. A C₁-C₄ straight chained or branched alkyl group isalso referred to as a “lower alkyl” group.

In any of the preceding embodiments, a C₁-C₄ alkoxy-substituted groupmay include one or more alkoxy substituents such as one, two or threemethoxy groups or a methoxy group and an ethoxy group, for example.Exemplary C₁-C₄ alkoxy substituents include methoxy, ethoxy, isopropoxy,and tert-butoxy.

In any of the preceding embodiments, a hydroxy-substituted group mayinclude one or more hydroxy substituents, such as two or three hydroxygroups.

A “halogen” refers to F, Cl, Br or I.

A “halogen-substitution” or “halo” substitution designates replacementof one or more hydrogens with F, Cl, Br or I.

In one aspect, the term haloalkyl is defined as any alkyl radical havingone or more hydrogen atoms replaced by a halogen atom. In any of thepreceding embodiments, a “halo-substituted” group includes from one halosubstituent up to perhalo substitution.

Exemplary halo-substituted C₁-C₄ alkyl includes CFH₂, CClH₂, CBrH₂,CF₂H, CCl₂H, CBr₂H, CF₃, CCl₃, CBr₃, CH₂CH₂F, CH₂CH₂Cl, CH₂CH₂Br,CH₂CHF₂, CHFCH₃, CHClCH₃, CHBrCH₃, CF₂CHF₂, CF₂CHCl₂, CF₂CHBr₂,CH(CF₃)₂, and C(CF₃)₃. Perhalo-substituted C₁-C₄ alkyl, for example,includes CF₃, CCl₃, CBr₃, CF₂CF₃, CCl₂CF₃ and CBr₂CF₃.

The terms “alkenyl” (“alkene”) and “alkynyl” (“alkyne”) refer tounsaturated aliphatic groups analogous in length and possiblesubstitution to the alkyl groups described above, but that contain atleast one double or triple bond respectively.

In any of the preceding embodiments, a “carbocycle” group may refer to amonocyclic carbocycle embodiment and/or a polycyclic carbocycleembodiment, such as a fused, bridged or bicyclic carbocycle embodiment.“Carbocycle” groups of the invention may further refer to an aromaticcarbocycle embodiment and/or a non-aromatic carbocycle embodiment, or,in the case of polycyclic embodiments, a carbocycle having both one ormore aromatic rings and/or one or more non-aromatic rings. Polycycliccarbocycle embodiments may be a bicyclic ring, a fused ring or a bridgedbicycle. Non-limiting exemplary carbocycles include phenyl, cyclohexane,cyclopentane, or cyclohexene, amantadine, cyclopentane, cyclohexane,bicyclo[2.2.1]heptane, 1,5-cyclooctadiene,1,2,3,4-tetrahydronaphthalene, bicyclo[4.2.0]oct-3-ene, naphthalene,adamantane, decalin, naphthalene, 1,2,3,4-tetrahydronaphthalene,norbornane, decalin, spiropentane, memantine, biperiden, rimantadine,camphor, cholesterol, 4-phenylcyclohexanol, bicyclo[4.2.0]octane,memantine and 4,5,6,7-tetrahydro-1H-indene and bicyclo[4.1.0]hept-3-ene.

In any of the preceding embodiments, a “heterocycle” group may refer toa monocyclic heterocycle embodiment and/or a polycyclic heterocyclicembodiment, such as a fused, bridged or bicyclic heterocycle embodiment.“Heterocycle” groups of the invention may further refer to an aromaticheterocycle embodiment and/or a non-aromatic heterocycle embodiment, or,in the case of polycyclic embodiments, a heterocycle having both one ormore aromatic rings and/or one or more non-aromatic rings. Polycyclicheterocycle embodiments may be a bicyclic ring, a fused ring or abridged bicycle. Non-limiting exemplary heterocycles include pyridyl,pyrrolidine, piperidine, piperazine, pyrrolidine, morpholine,pyrimidine, benzofuran, indole, quinoline, lactones, lactams,benzodiazepine, indole, quinoline, purine, adenine, guanine,4,5,6,7-tetrahydrobenzo[d]thiazole, hexamine and methenamine.

“Alkenyl” refers to an unsaturated hydrocarbon chain having thespecified number of member carbon atoms and having one or morecarbon-carbon double bonds within the chain. For example, C2-C6 alkenylrefers to an alkenyl group having from 2 to 6 member carbon atoms. Incertain embodiments, alkenyl groups have one carbon-carbon double bondwithin the chain. In other embodiments, alkenyl groups have more thanone carbon-carbon double bond within the chain. Alkenyl groups may beoptionally substituted with one or more substituents as defined herein.Alkenyl groups may be straight or branched. Representative branchedalkenyl groups have one, two, or three branches. Alkenyl includesethylenyl, propenyl, butenyl, pentenyl, and hexenyl.

“Alkoxy” refers to an alkyl moiety attached through an oxygen bridge(i.e. a —O-C1-C6 alkyl group wherein C1-C6 is defined herein). Examplesof such groups include methoxy, ethoxy, propoxy, butoxy, pentoxy andhexoxy.

“Alkynyl” refers to an unsaturated hydrocarbon chain having thespecified number of member carbon atoms and having one or morecarbon-carbon triple bonds within the chain. For example, C2-C6 alkynylrefers to an alkynyl group having from 2 to 6 member atoms. In certainembodiments alkynyl groups have one carbon-carbon triple bond within thechain. In other embodiments, alkynyl groups have more than onecarbon-carbon triple bond within the chain. For the sake of clarity,unsaturated hydrocarbon chains having one or more carbon-carbon triplebond within the chain and one or more carbon-carbon double bond withinthe chain are referred to as alkynyl groups. Alkynyl groups may beoptionally substituted with one or more substituents as defined herein.Representative branched alkynyl groups have one, two, or three branches.Alkynyl includes ethynyl, propynyl, butynyl, pentynyl, and hexynyl.

The term “aromatic carbocycle” refers to an aromatic hydrocarbon ringsystem containing at least one aromatic ring. The ring may be fused orotherwise attached to other aromatic carbocyclic rings or non-aromaticcarbocyclic rings. Examples of aromatic carbocycle groups includecarbocyclic aromatic groups such as phenyl, naphthyl, and anthracyl.

“Azabicyclo” refers to a bicyclic molecule that contains a nitrogen atomin the ring skeleton. The two rings of the bicycle may be fused at twomutually bonded atoms, e.g., indole, across a sequence of atoms, e.g.,azabicyclo[2.2.1]heptane, or joined at a single atom, e.g., spirocycle.

“Bicycle” or “bicyclic” refers to a two-ring system in which one, two orthree or more atoms are shared between the two rings. Bicycle includesfused bicycles in which two adjacent atoms are shared by each of the tworings, e.g., decalin, indole. Bicycle also includes spiro bicycles inwhich two rings share a single atom, e.g., spiro[2.2]pentane,1-oxa-6-azaspiro[3.4]octane. Bicycle further includes bridged bicyclesin which at least three atoms are shared between two rings, e.g.,norbornane.

“Bridged bicycle” compounds are bicyclic ring systems in which at leastthree atoms are shared by both rings of the system, i.e., they includeat least one bridge of one or more atoms connecting two bridgeheadatoms. Bridged azabicyclo refers to a bridged bicyclic molecule thatcontains a nitrogen atom in at least one of the rings.

The term “Boc” refers to a tert-butyloxycarbonyl group (a common amineprotecting group).

The terms “carbocycle”, and “carbocyclic”, as used herein, refers to asaturated or unsaturated ring in which each atom of the ring is carbon.The term carbocycle includes both aromatic carbocycles and non-aromaticcarbocycles. Non-aromatic carbocycles include both cycloalkane rings, inwhich all carbon atoms are saturated, and cycloalkene rings, whichcontain at least one double bond. “Carbocycle” includes 5-7 memberedmonocyclic and 8-12 membered bicyclic rings. Each ring of a bicycliccarbocycle may be selected fromnon-aromatic and aromatic rings.Carbocycle includes bicyclic molecules in which one, two or three ormore atoms are shared between the two rings. The term “fused carbocycle”refers to a bicyclic carbocycle in which each of the rings shares twoadjacent atoms with the other ring. Each ring of a fused carbocycle maybe selected fromnon-aromaticaromatic rings. In an exemplary embodiment,an aromatic ring, e.g., phenyl, may be fused to a non-aromatic oraromatic ring, e.g., cyclohexane, cyclopentane, or cyclohexene. Anycombination of non-aromatic and aromatic bicyclic rings, as valencepermits, is included in the definition of carbocyclic. Exemplary“carbocycles” include cyclopentane, cyclohexane, bicyclo[2.2.1]heptane,1,5-cyclooctadiene, 1,2,3,4-tetrahydronaphthalene,bicyclo[4.2.0]oct-3-ene, naphthalene and adamantane. Exemplary fusedcarbocycles include decalin, naphthalene, 1,2,3,4-tetrahydronaphthalene,bicyclo[4.2.0]octane, 4,5,6,7-tetrahydro-1H-indene andbicyclo[4.1.0]hept-3-ene. “Carbocycles” may be substituted at any one ormore positions capable of bearing a hydrogen atom.

A “cycloalkyl” group is a cyclic hydrocarbon ring having the specifiednumber of member carbon atoms which is completely saturated(non-aromatic). Typically, a cycloalkyl group has from 3 to about 10carbon atoms, more typically 3 to 8 carbon atoms unless otherwisedefined. Cycloalkyl groups are monocyclic ring systems. For example,C3-C6 cycloalkyl refers to a cycloalkyl group having from 3 to 6 memberatoms. Cycloalkyl groups may be optionally substituted with one or moresubstituents as defined herein. Cycloalkyl includes cyclopropyl,cyclobutyl, cyclopentyl, and cyclohexyl.

A “cycloalkenyl” group is a cyclic hydrocarbon ring containing one ormore double bonds within the ring. For example, C3-C6 cycloalkenylrefers to a cycloalkenyl group having from 3 to 6 member carbon atoms.In certain embodiments, cycloalkenyl groups have one carbon-carbondouble bond within the ring. In other embodiments, cycloalkenyl groupshave more than one carbon-carbon double bonds within the ring.Cycloalkenyl rings are not aromatic. Cycloalkenyl groups are monocyclicring systems. Cycloalkenyl groups may be optionally substituted with oneor more substituents as defined herein. Cycloalkenyl includescyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, andcyclohexadienyl.

“Aryl” refers to an aromatic hydrocarbon ring system. Aryl groups aremonocyclic ring systems or bicyclic ring systems. Monocyclic aryl ringrefers to phenyl. Bicyclic aryl rings refer to napthyl and to ringswherein phenyl is fused to a cycloalkyl or cycloalkenyl ring having 5,6, or 7 member carbon atoms. Aryl groups may be optionally substitutedwith one or more substituents as defined herein.

The term “heteroaryl” or “aromatic heterocycle” includes substituted orunsubstituted aromatic single ring structures, preferably 5- to7-membered rings, more preferably 5- to 6-membered rings, whose ringstructures include at least one heteroatom, preferably one to fourheteroatoms, more preferably one or two heteroatoms. The term“heteroaryl” also includes ring systems having one or two rings whereinat least one of the rings is heteroaromatic, e.g., the other cyclicrings can be cycloalkyl, cycloalkenyl, cycloalkynyl, aromaticcarbocycle, heteroaryl, and/or heterocyclyl. Heteroaryl groups include,for example, pyrrole, furan, thiophene, imidazole, oxazole, thiazole,pyrazole, pyridine, pyrazine, pyridazine, and pyrimidine.

The terms “heterocycle”, and “heterocyclic”, as used herein, refers to anon-aromatic or aromatic ring comprising one or more heteroatomsselected from, for example, N, O, B and S atoms, preferably N, O, or S.The term “heterocycle” includes both “aromatic heterocycles” and“non-aromatic heterocycles.” Heterocycles include 4-7 memberedmonocyclic and 8-12 membered bicyclic rings. Heterocycle includesbicyclic molecules in which one, two or three or more atoms are sharedbetween the two rings. Each ring of a bicyclic heterocycle may beselected fromnon-aromatic and aromatic rings. The term “fusedheterocycle” refers to a bicyclic heterocycle in which each of the ringsshares two adjacent atoms with the other ring. Each ring of a fusedheterocycle may be selected fromnon-aromatic and aromatic rings. In anexemplary embodiment, an aromatic ring, e.g., pyridyl, may be fused to anon-aromatic or aromatic ring, e.g., cyclohexane, cyclopentane,pyrrolidine, 2,3-dihydrofuran or cyclohexene. “Heterocycle” groupsinclude, for example, piperidine, piperazine, pyrrolidine, morpholine,pyrimidine, benzofuran, indole, quinoline, lactones, and lactams.Exemplary “fused heterocycles” include benzodiazepine, indole,quinoline, purine, and 4,5,6,7-tetrahydrobenzo[d]thiazole.“Heterocycles” may be substituted at any one or more positions capableof bearing a hydrogen atom.

“Monocyclic rings” include 5-7 membered aromatic carbocycle orheteroaryl, 3-7 membered cycloalkyl or cycloalkenyl, and 5-7 memberednon-aromatic heterocyclyl. Exemplary monocyclic groups includesubstituted or unsubstituted heterocycles or carbocycles such asthiazolyl, oxazolyl, oxazinyl, thiazinyl, dithianyl, dioxanyl,isoxazolyl, isothiazolyl, triazolyl, furanyl, tetrahydrofuranyl,dihydrofuranyl, pyranyl, tetrazolyl, pyrazolyl, pyrazinyl, pyridazinyl,imidazolyl, pyridinyl, pyrrolyl, dihydropyrrolyl, pyrrolidinyl,piperidinyl, piperazinyl, pyrimidinyl, morpholinyl,tetrahydrothiophenyl, thiophenyl, cyclohexyl, cyclopentyl, cyclopropyl,cyclobutyl, cycloheptanyl, azetidinyl, oxetanyl, thiiranyl, oxiranyl,aziridinyl, and thiomorpholinyl.

-   -   “Member atoms” refers to the atom or atoms that form a chain or        ring. Where more than one member atom is present in a chain and        within a ring, each member atom is covalently bound to an        adjacent member atom in the chain or ring. Atoms that make up a        substituent group on a chain or ring are not member atoms in the        chain or ring.        -   “Optionally substituted” indicates that a group, such as            alkyl, alkenyl, alkynyl, aryl, cycloalkyl, cycloalkenyl,            heterocycloalkyl, or heteroaryl, may be unsubstituted, or            the group may be substituted with one or more substituents            as defined herein.

As used herein, “substituted” means substituting a hydrogen atom in astructure with an atom or molecule other than hydrogen. “Substituted” inreference to a group indicates that one or more hydrogen atoms attachedto a member atom within the group is replaced with a substituentselected from the group of defined substituents. A substitutable atomsuch as a “substitutable nitrogen” is an atom that bears a hydrogen atomin at least one resonance form. The hydrogen atom may be substituted foranother atom or group such as a CH₃ or an OH group. For example, thenitrogen in a piperidine molecule is substitutable if the nitrogen isbound to a hydrogen atom. If, for example, the nitrogen of a piperidineis bound to an atom other than hydrogen, the nitrogen is notsubstitutable. An atom that is not capable of bearing a hydrogen atom inany resonance form is not substitutable. It should be understood thatthe term “substituted” includes the implicit provision that suchsubstitution be in accordance with the permitted valence of thesubstituted atom and the substituent, and that the substitution resultsin a stable compound (i.e. one that does not spontaneously undergotransformation such as by hydrolysis, rearrangement, cyclization, orelimination, and that is sufficiently robust to survive isolation from areaction mixture). When it is stated that a group may contain one ormore substituents, one or more (as appropriate) member atom within thegroup may be substituted. In addition, a single member atom within thegroup may be substituted with more than one substituent as long as suchsubstitution is in accordance with the permitted valence of the atom.Suitable substituents are defined herein for each substituted oroptionally substituted group.

Combinations of substituents and variables envisioned by this inventionare only those that result in the formation of stable compounds. As usedherein, the term “stable” refers to compounds that possess stabilitysufficient to allow manufacture and that maintain the integrity of thecompound for a sufficient period of time to be useful for the purposesdetailed herein.

Deutrated Compounds

The compounds disclosed herein also include partially and fullydeuterated variants. In certain embodiments, deuterated variants may beused for kinetic studies. One of skill in the art can select the sitesat which such deuterium atoms are present.

The invention also includes various deuterated forms of the compounds ofFormulas (I) or pharmaceutically acceptable salts thereof. Eachavailable hydrogen atom attached to a carbon atom may be independentlyreplaced with a deuterium atom.

A person of ordinary skill in the art will know how to synthesizedeuterated forms of the compounds of Formulas (I) to (II) of the presentinvention. For example, deuterated materials, such as alkyl groups maybe prepared by conventional techniques (see for example: methyl-d₃-amineavailable from Aldrich Chemical Co., Milwaukee, Wis., Cat. No. 489,689-2).

Isotopes

The subject invention also includes isotopically-labeled compounds whichare identical to those recited in Formulas (I) and (II) but for the factthat one or more atoms are replaced by an atom having an atomic mass ormass number different from the atomic mass or mass number most commonlyfound in nature. Examples of isotopes that can be incorporated intocompounds of the invention include isotopes of hydrogen, carbon,nitrogen, oxygen, fluorine, iodine and chlorine such as ³H, ¹¹C, ¹⁴C,¹⁸F, ¹²³I or ¹²⁵I.

Compounds of the present invention and pharmaceutically acceptable saltsof said compounds that contain the aforementioned isotopes and/or otherisotopes of other atoms are within the scope of the present invention.Isotopically labeled compounds of the present invention, for examplethose into which radioactive isotopes such as ³H or ¹⁴C have beenincorporated, are useful in drug and/or substrate tissue distributionassays. Tritiated, ie. ³H, and carbon-14, ie. ¹⁴C, isotopes areparticularly preferred for their ease of preparation and detectability.¹¹C and ¹⁸F isotopes are particularly useful in PET (positron emissiontomography).

Purity

Because the compounds of the present invention are intended for use inpharmaceutical compositions it will readily be understood that they areeach preferably provided in substantially pure form, for example atleast 60% pure, more suitably at least 75% pure and preferably at least85%, especially at least 98% pure (% are on a weight for weight basis).Impure preparations of the compounds may be used for preparing the morepure forms used in the pharmaceutical compositions.

Salts

In certain embodiments, compounds according to Formula I or apharmaceutically acceptable salt thereof may contain an acidicfunctional group. In certain other embodiments, compounds according toFormula I may contain a basic functional group. Thus, the skilledartisan will appreciate that salts of the compounds according to FormulaI may be prepared. Indeed, in certain embodiments of the invention,salts of the compounds according to Formula I may be preferred over therespective free base or free acid because, for example, such salts mayimpart greater stability or solubility to the molecule therebyfacilitating formulation into a dosage form.

Because of their potential use in medicine, the salts of the compoundsof Formulas (I) are suitably pharmaceutically acceptable salts. Suitablepharmaceutically acceptable salts include those described by Berge,Bighley and Monkhouse J.Pharm.Sci (1977) 66, pp 1-19.

Also included in the present invention are salts, particularlypharmaceutically acceptable salts, of the compounds described herein.The compounds of the present invention that possess a sufficientlyacidic, a sufficiently basic, or both functional groups, can react withany of a number of inorganic bases, and inorganic and organic acids, toform a salt. Alternatively, compounds that are inherently charged, suchas those with quaternary nitrogen, can form a salt with an appropriatecounterion (e.g., a halide such as bromide, chloride, or fluoride,particularly bromide).

Acids commonly employed to form acid addition salts are inorganic acidssuch as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuricacid, phosphoric acid, and the like, and organic acids such asp-toluenesulfonic acid, methanesulfonic acid, oxalic acid,p-bromophenyl-sulfonic acid, carbonic acid, succinic acid, citric acid,benzoic acid, acetic acid, and the like. Examples of such salts includethe sulfate, pyrosulfate, bisulfate, sulfite, bisulfate, phosphate,monohydrogenphosphate, dihydrogenphosphate, metaphosphate,pyrophosphate, chloride, bromide, iodide, acetate, propionate,decanoate, caprylate, acrylate, formate, isobutyrate, caproate,heptanoate, propiolate, oxalate, malonate, succinate, suberate,sebacate, fumarate, maleate, butyne-1,4-dioate, hexyne-1,6-dioate,benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate,hydroxybenzoate, methoxybenzoate, phthalate, sulfonate, xylenesulfonate,phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate,gamma-hydroxybutyrate, glycolate, tartrate, methanesulfonate,propanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate,mandelate, and the like.

Base addition salts include those derived from inorganic bases, such asammonium or alkali or alkaline earth metal hydroxides, carbonates,bicarbonates, and the like. Such bases useful in preparing the salts ofthis invention thus include sodium hydroxide, potassium hydroxide,ammonium hydroxide, potassium carbonate, and the like.

“Enantiomeric excess” or “ee” is the excess of one enantiomer over theother expressed as a percentage. As a result, since both enantiomers arepresent in equal amounts in a racemic mixture, the enantiomeric excessis zero (0% ee). However, if one enantiomer was enriched such that itconstitutes 95% of the product, then the enantiomeric excess would be90% ee (the amount of the enriched enantiomer, 95%, minus the amount ofthe other enantiomer, 5%).

“Enantiomerically enriched” refers to products whose enantiomeric excessis greater than zero. For example, enantiomerically enriched refers toproducts whose enantiomeric excess is greater than 50% ee, greater than75% ee, or greater than 90% ee.

“Enantiomerically pure” refers to products whose enantiomeric excess is99% ee or greater.

“Pharmaceutically acceptable” refers to those compounds, materials,compositions, and dosage forms which are, within the scope of soundmedical judgment, suitable for use in contact with the tissues of humanbeings or animals without excessive toxicity, irritation, or otherproblem or complication, commensurate with a reasonable benefit/riskratio.The compounds according to Formula (I) or a pharmaceutically acceptablesalt thereof, may contain one or more asymmetric centers (also referredto as a chiral center) and may, therefore, exist as individualenantiomers, diastereomers, or other stereoisomeric forms, or asmixtures thereof.Chiral centers, such as chiral carbon atoms, may also be present in asubstituent such as an alkyl group. Where the stereochemistry of achiral center present in Formula I, or in any chemical structureillustrated herein, is not specified, the structure is intended toencompass all individual stereoisomers and all mixtures thereof.Thus, compounds according to Formula (I) or pharmaceutically acceptablesalts thereof, containing one or more chiral centers may be used asracemic mixtures, diastereomeric mixtures, enantiomerically enrichedmixtures, diastereomerically enriched mixtures, or as enantiomericallyand diastereomerically pure individual stereoisomers.

Individual stereoisomers of a compound according to Formula (I) or apharmaceutically acceptable salt thereof which contain one or moreasymmetric centers may be resolved by methods known to those skilled inthe art. For example, such resolution may be carried out (1) byformation of diastereoisomeric salts, complexes or other derivatives;(2) by selective reaction with a stereoisomer-specific reagent, forexample by enzymatic oxidation or reduction; or (3) by gas-liquid orliquid chromatography in a chiral environment, for example, on a chiralsupport such as silica with a bound chiral ligand or in the presence ofa chiral solvent. The skilled artisan will appreciate that where thedesired stereoisomer is converted into a diastereomeric salt, complex orderivative, a further step is required to liberate the desired form.Alternatively, specific stereoisomers may be synthesized by asymmetricsynthesis using optically active reagents, substrates, catalysts orsolvents, or by converting one enantiomer to the other by asymmetrictransformation.

When a disclosed compound or its salt is named or depicted by structure,it is to be understood that the compound or salt, including solvates(particularly, hydrates) thereof, may exist in crystalline forms,non-crystalline forms or a mixture thereof. The compound or salt, orsolvates (particularly, hydrates) thereof, may also exhibit polymorphism(i.e. the capacity to occur in different crystalline forms). Thesedifferent crystalline forms are typically known as “polymorphs.”

In light of this, salt forms of the present invention (i.e., which mayinclude different polymorphs, anhydrous forms, solvates, or hydratesthereof) may exhibit characteristic polymorphism. As conventionallyunderstood in the art, polymorphism is defined as an ability of acompound to crystallize as more than one distinct crystalline or“polymorphic” species. A polymorph is defined as a solid crystallinephase of a compound with at least two different arrangements orpolymorphic forms of that compound molecule in the solid state.

Polymorphic forms of any given compound, including those of the presentinvention, are defined by the same chemical formula or composition andare as distinct in chemical structure as crystalline structures of twodifferent chemical compounds. Such compounds may differ in packing,geometrical arrangement of respective crystalline lattices, etc.

It is to be understood that when named or depicted by structure, thedisclosed compound, or solvates (particularly, hydrates) thereof, alsoinclude all polymorphs thereof. Polymorphs have the same chemicalcomposition but differ in packing, geometrical arrangement, and otherdescriptive properties of the crystalline solid state.

In light of the foregoing, chemical and/or physical properties orcharacteristics vary with each distinct polymorphic form, which mayinclude variations in solubility, melting point, density, hardness,crystal shape, optical and electrical properties, vapor pressure,stability, etc.

Solvates and/or hydrates of crystalline salt forms of the presentinvention also may be formed when solvent molecules are incorporatedinto the crystalline lattice structure of the compound molecule duringthe crystallization process. For example, solvate forms of the presentinvention may incorporate nonaqueous solvents such as methanol and thelike as described herein below. Hydrate forms are solvate forms, whichincorporate water as a solvent into a crystalline lattice.

Anhydrous with respect to solid state polymorphism refers to acrystalline structure that does not contain a repeating, crystallinesolvent in the lattice. However, crystalline materials can be porous andmay exhibit reversible surface adsorption of water.

Terms and Definitions Section 2 1. Definitions

As used herein, the following terms and phrases shall have the meaningsset forth below. Unless defined otherwise, all technical and scientificterms used herein have the same meaning as commonly understood to one ofordinary skill in the art.

The term “agent” is used herein to denote a chemical compound, a mixtureof chemical compounds, a biological macromolecule (such as a nucleicacid, an antibody, a protein or portion thereof, e.g., a peptide), or anextract made from biological materials such as bacteria, plants, fungi,or animal (particularly mammalian) cells or tissues.

The term “bioavailable”, when referring to a compound, is art-recognizedand refers to a form of a compound that allows for all or a portion ofthe amount of compound administered to be absorbed by, incorporatedinto, or otherwise physiologically available to a subject or patient towhom it is administered.

“Biologically active portion of a sirtuin” refers to a portion of asirtuin protein having a biological activity, such as the ability todeacetylate (“catalytically active”). Catalytically active portions of asirtuin may comprise the core domain of sirtuins. Catalytically activeportions of SIRT1 having GenBank Accession No. NP_036370 that encompassthe NAD⁺ binding domain and the substrate binding domain, for example,may include without limitation, amino acids 240-664 or 240-505 ofGenBank Accession No. NP_036370, which are encoded by the polynucleotideof GenBank Accession No. NM_012238. Therefore, this region is sometimesreferred to as the core domain. Other catalytically active portions ofSIRT1, also sometimes referred to as core domains, include about aminoacids 261 to 447 of GenBank Accession No. NP_036370, which are encodedby nucleotides 834 to 1394 of GenBank Accession No. NM_012238; aboutamino acids 242 to 493 of GenBank Accession No. NP_036370, which areencoded by nucleotides 777 to 1532 of GenBank Accession No. NM_012238;or about amino acids 254 to 495 of GenBank Accession No. NP_036370,which are encoded by nucleotides 813 to 1538 of GenBank Accession No.NM_012238. Another “biologically active” portion of SIRT1 is amino acids62-293 or 183-225 of GenBank Accession No. NP_036370, which comprise adomain N-terminal to the core domain that is important to the compoundbinding site.

The term “companion animals” refers to cats and dogs. As used herein,the term “dog(s)” denotes any member of the species Canis familiaris, ofwhich there are a large number of different breeds. The term “cat(s)”refers to a feline animal including domestic cats and other members ofthe family Felidae, genus Felis.

“Diabetes” refers to high blood sugar or ketoacidosis, as well aschronic, general metabolic abnormalities arising from a prolonged highblood sugar status or a decrease in glucose tolerance. “Diabetes”encompasses both the type I and type II (Non Insulin Dependent DiabetesMellitus or NIDDM) forms of the disease. The risk factors for diabetesinclude the following factors: waistline of more than 40 inches for menor 35 inches for women, blood pressure of 130/85 mmHg or higher,triglycerides above 150 mg/dl, fasting blood glucose greater than 100mg/dl or high-density lipoprotein of less than 40 mg/dl in men or 50mg/dl in women.

The term “ED₅₀” refers to the art-recognized measure of effective dose.In certain embodiments, ED₅₀ means the dose of a drug which produces 50%of its maximum response or effect, or alternatively, the dose whichproduces a pre-determined response in 50% of test subjects orpreparations, such as isolated tissue or cells. The term “LD₅₀” refersto the art-recognized measure of lethal dose. In certain embodiments,LD₅₀ means the dose of a drug which is lethal in 50% of test subjects.The term “therapeutic index” is an art-recognized term which refers tothe therapeutic index of a drug, defined as LD₅₀/ED₅₀.

The term “hyperinsulinemia” refers to a state in an individual in whichthe level of insulin in the blood is higher than normal.

The term “insulin resistance” refers to a state in which a normal amountof insulin produces a subnormal biologic response relative to thebiological response in a subject that does not have insulin resistance.

An “insulin resistance disorder,” as discussed herein, refers to anydisease or condition that is caused by or contributed to by insulinresistance. Examples include: diabetes, obesity, metabolic syndrome,insulin-resistance syndromes, syndrome X, insulin resistance, high bloodpressure, hypertension, high blood cholesterol, dyslipidemia,hyperlipidemia, atherosclerotic disease including stroke, coronaryartery disease or myocardial infarction, hyperglycemia, hyperinsulinemiaand/or hyperproinsulinemia, impaired glucose tolerance, delayed insulinrelease, diabetic complications, including coronary heart disease,angina pectoris, congestive heart failure, stroke, cognitive functionsin dementia, retinopathy, peripheral neuropathy, nephropathy,glomerulonephritis, glomerulosclerosis, nephrotic syndrome, hypertensivenephrosclerosis, some types of cancer (such as endometrial, breast,prostate, and colon), complications of pregnancy, poor femalereproductive health (such as menstrual irregularities, infertility,irregular ovulation, polycystic ovarian syndrome (PCOS)), lipodystrophy,cholesterol-related disorders, such as gallstones, cholecystitis andcholelithiasis, gout, obstructive sleep apnea and respiratory problems,osteoarthritis, and bone loss, e.g., osteoporosis in particular.

The term “livestock animals” refers to domesticated quadrupeds, whichincludes those being raised for meat and various byproducts, e.g., abovine animal including cattle and other members of the genus Bos, aporcine animal including domestic swine and other members of the genusSus, an ovine animal including sheep and other members of the genusOvis, domestic goats and other members of the genus Capra; domesticatedquadrupeds being raised for specialized tasks such as use as a beast ofburden, e.g., an equine animal including domestic horses and othermembers of the family Equidae, genus Equus.

The term “mammal” is known in the art, and exemplary mammals includehumans, primates, livestock animals (including bovines, porcines, etc.),companion animals (e.g., canines, felines, etc.) and rodents (e.g., miceand rats).

“Obese” individuals or individuals suffering from obesity are generallyindividuals having a body mass index (BMI) of at least 25 or greater.Obesity may or may not be associated with insulin resistance.

The terms “parenteral administration” and “administered parenterally”are art-recognized and refer to modes of administration other thanenteral and topical administration, usually by injection, and includes,without limitation, intravenous, intramuscular, intraarterial,intrathecal, intracapsular, intraorbital, intracardiac, intradermal,intraperitoneal, transtracheal, subcutaneous, subcuticular,intra-articular, subcapsular, subarachnoid, intraspinal, andintrasternal injection and infusion.

A “patient”, “subject”, “individual” or “host” refers to either a humanor a non-human animal.

The term “pharmaceutically acceptable carrier” is art-recognized andrefers to a pharmaceutically-acceptable material, composition orvehicle, such as a liquid or solid filler, diluent, excipient, solventor encapsulating material, involved in carrying or transporting anysubject composition or component thereof. Each carrier must be“acceptable” in the sense of being compatible with the subjectcomposition and its components and not injurious to the patient. Someexamples of materials which may serve as pharmaceutically acceptablecarriers include: (1) sugars, such as lactose, glucose and sucrose; (2)starches, such as corn starch and potato starch; (3) cellulose, and itsderivatives, such as sodium carboxymethyl cellulose, ethyl cellulose andcellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7)talc; (8) excipients, such as cocoa butter and suppository waxes; (9)oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil,olive oil, corn oil and soybean oil; (10) glycols, such as propyleneglycol; (11) polyols, such as glycerin, sorbitol, mannitol andpolyethylene glycol; (12) esters, such as ethyl oleate and ethyllaurate; (13) agar; (14) buffering agents, such as magnesium hydroxideand aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17)isotonic saline; (18) Ringer's solution; (19) ethyl alcohol; (20)phosphate buffer solutions; and (21) other non-toxic compatiblesubstances employed in pharmaceutical formulations.

The term “preventing” is art-recognized, and when used in relation to acondition, such as a local recurrence (e.g., pain), a disease such ascancer, a syndrome complex such as heart failure or any other medicalcondition, is well understood in the art, and includes administration ofa composition which reduces the frequency of, or delays the onset of,symptoms of a medical condition in a subject relative to a subject whichdoes not receive the composition. Thus, prevention of cancer includes,for example, reducing the number of detectable cancerous growths in apopulation of patients receiving a prophylactic treatment relative to anuntreated control population, and/or delaying the appearance ofdetectable cancerous growths in a treated population versus an untreatedcontrol population, e.g., by a statistically and/or clinicallysignificant amount. Prevention of an infection includes, for example,reducing the number of diagnoses of the infection in a treatedpopulation versus an untreated control population, and/or delaying theonset of symptoms of the infection in a treated population versus anuntreated control population. Prevention of pain includes, for example,reducing the magnitude of, or alternatively delaying, pain sensationsexperienced by subjects in a treated population versus an untreatedcontrol population.

The term “prophylactic” or “therapeutic” treatment is art-recognized andrefers to administration of a drug to a host. If it is administeredprior to clinical manifestation of the unwanted condition (e.g., diseaseor other unwanted state of the host animal) then the treatment isprophylactic, i.e., it protects the host against developing the unwantedcondition, whereas if administered after manifestation of the unwantedcondition, the treatment is therapeutic (i.e., it is intended todiminish, ameliorate or maintain the existing unwanted condition or sideeffects therefrom).

The term “pyrogen-free”, with reference to a composition, refers to acomposition that does not contain a pyrogen in an amount that would leadto an adverse effect (e.g., irritation, fever, inflammation, diarrhea,respiratory distress, endotoxic shock, etc.) in a subject to which thecomposition has been administered. For example, the term is meant toencompass compositions that are free of, or substantially free of, anendotoxin such as, for example, a lipopolysaccharide (LPS).

“Replicative lifespan” of a cell refers to the number of daughter cellsproduced by an individual “mother cell.” “Chronological aging” or“chronological lifespan,” on the other hand, refers to the length oftime a population of non-dividing cells remains viable when deprived ofnutrients. “Increasing the lifespan of a cell” or “extending thelifespan of a cell,” as applied to cells or organisms, refers toincreasing the number of daughter cells produced by one cell; increasingthe ability of cells or organisms to cope with stresses and combatdamage, e.g., to DNA, proteins; and/or increasing the ability of cellsor organisms to survive and exist in a living state for longer under aparticular condition, e.g., stress (for example, heatshock, osmoticstress, high energy radiation, chemically-induced stress, DNA damage,inadequate salt level, inadequate nitrogen level, or inadequate nutrientlevel). Lifespan can be increased by at least about 10%, 20%, 30%, 40%,50%, 60% or between 20% and 70%, 30% and 60%, 40% and 60% or more usingmethods described herein.

“Sirtuin-modulating compound” refers to a compound that increases thelevel of a sirtuin protein and/or increases at least one activity of asirtuin protein. In an exemplary embodiment, a sirtuin-modulatingcompound may increase at least one biological activity of a sirtuinprotein by at least about 10%, 25%, 50%, 75%, 100%, or more. Exemplarybiological activities of sirtuin proteins include deacetylation, e.g.,of histones and p53; extending lifespan; increasing genomic stability;silencing transcription; and controlling the segregation of oxidizedproteins between mother and daughter cells.

proteins include deacetylation, e.g., of an acetylated peptidesubstrate.

“Sirtuin protein” refers to a member of the sirtuin deacetylase proteinfamily, or preferably to the sir2 family, which include yeast Sir2(GenBank Accession No. P53685), C. elegans Sir-2.1 (GenBank AccessionNo. NP 501912), and human SIRT1 (GenBank Accession No. NM_012238 andNP_036370 (or AF083106)) and SIRT2 (GenBank Accession No. NM_012237,NM_030593, NP_036369, NP_085096, and AF083107) proteins. Other familymembers include the four additional yeast Sir2-like genes termed “HSTgenes” (homologues of Sir two) HST1, HST2, HST3 and HST4, and the fiveother human homologues hSIRT3, hSIRT4, hSIRT5, hSIRT6 and hSIRT7(Brachmann et al. (1995) Genes Dev. 9:2888 and Frye et al. (1999) BBRC260:273).

“SIRT1 protein” refers to a member of the sir2 family of sirtuindeacetylases. In certain embodiments, a SIRT1 protein includes yeastSir2 (GenBank Accession No. P53685), C. elegans Sir-2.1 (GenBankAccession No. NP_501912), human SIRT1 (GenBank Accession No. NM_012238or NP_036370 (or AF083106)), mouse SIRT1 (GenBank Accession No.NM_019812 or NP_062786), and equivalents and fragments thereof. Inanother embodiment, a SIRT1 protein includes a polypeptide comprising asequence consisting of, or consisting essentially of, the amino acidsequence set forth in GenBank Accession Nos. NP_036370, NP_501912,NP_085096, NP_036369, or P53685. SIRT1 proteins include polypeptidescomprising all or a portion of the amino acid sequence set forth inGenBank Accession Nos. NP_036370, NP_501912, NP_085096, NP_036369, orP53685; the amino acid sequence set forth in GenBank Accession Nos.NP_036370, NP_501912, NP_085096, NP_036369, or P53685 with 1 to about 2,3, 5, 7, 10, 15, 20, 30, 50, 75 or more conservative amino acidsubstitutions; an amino acid sequence that is at least 60%, 70%, 80%,90%, 95%, 96%, 97%, 98%, or 99% identical to GenBank Accession Nos.NP_036370, NP_501912, NP_085096, NP_036369, or P53685, and functionalfragments thereof. Polypeptides of the invention also include homologs(e.g., orthologs and paralogs), variants, or fragments, of GenBankAccession Nos. NP_036370, NP_501912, NP_085096, NP_036369, or P53685.

As used herein “SIRT2 protein”, “SIRT3 protein”, “SIRT4 protein”, SIRT5protein”, “SIRT6 protein”, and “SIRT7 protein” refer to other mammalian,e.g. human, sirtuin deacetylase proteins that are homologous to SIRT1protein, particularly in the approximately 275 amino acid conservedcatalytic domain. For example, “SIRT3 protein” refers to a member of thesirtuin deacetylase protein family that is homologous to SIRT1 protein.In certain embodiments, a SIRT3 protein includes human SIRT3 (GenBankAccession No. AAH01042, NP_036371, or NP_001017524) and mouse SIRT3(GenBank Accession No. NP_071878) proteins, and equivalents andfragments thereof. In certain embodiments, a SIRT4 protein includeshuman SIRT4 (GenBank Accession No. NM_012240 or NP_036372). In certainembodiments, a SIRT5 protein includes human SIRT5 (GenBank Accession No.NM_012241 or NP_036373). In certain embodiments, a SIRT6 proteinincludes human SIRT6 (GenBank Accession No. NM_016539 or NP_057623). Inanother embodiment, a SIRT3 protein includes a polypeptide comprising asequence consisting of, or consisting essentially of, the amino acidsequence set forth in GenBank Accession Nos. AAH01042, NP_036371,NP_001017524, or NP_071878. SIRT3 proteins include polypeptidescomprising all or a portion of the amino acid sequence set forth inGenBank Accession AAH01042, NP_036371, NP_001017524, or NP_071878; theamino acid sequence set forth in GenBank Accession Nos. AAH01042,NP_036371, NP_001017524, or NP_071878 with 1 to about 2, 3, 5, 7, 10,15, 20, 30, 50, 75 or more conservative amino acid substitutions; anamino acid sequence that is at least 60%, 70%, 80%, 90%, 95%, 96%, 97%,98%, or 99% identical to GenBank Accession Nos. AAH01042, NP_036371,NP_001017524, or NP_071878, and functional fragments thereof.Polypeptides of the invention also include homologs (e.g., orthologs andparalogs), variants, or fragments, of GenBank Accession Nos. AAH01042,NP_036371, NP_001017524, or NP_071878. In certain embodiments, a SIRT3protein includes a fragment of SIRT3 protein that is produced bycleavage with a mitochondrial matrix processing peptidase (MPP) and/or amitochondrial intermediate peptidase (MIP).

The term “steroisomer” as used herein is art-recognized and refers toany of two or more isomers that have the same molecular constitution anddiffer only in the three-dimensional arrangement of their atomicgroupings in space. When used herein to describe a compounds or genus ofcompounds, stereoisomer includes any portion of the compound or thecompound in its entirety. For example, diastereomers and enantiomers arestereoisomers.

The terms “systemic administration” and “administered systemically,” areart-recognized and refer to the administration of a subject composition,therapeutic or other material enterally or parenterally.

The term “tautomer” as used herein is art-recognized and refers to anyone of the possible alternative structures that may exist as a result oftautomerism, which refers to a form of constitutional isomerism in whicha structure may exist in two or more constitutional arrangements,particularly with respect to the position of hydrogens bonded to oxygen.When used herein to describe a compound or genus of compounds, it isfurther understood that a “tautomer” is readily interconvertible andexists in equilibrium. For example, keto and enol tautomers exist inproportions determined by the equilibrium position for any givencondition, or set of conditions:

The term “therapeutic agent” is art-recognized and refers to anybiologically, physiologically, or pharmacologically active substancethat acts locally or systemically in a subject. The term also means anysubstance intended for use in the diagnosis, cure, mitigation, treatmentor prevention of disease or in the enhancement of desirable physical ormental development and/or conditions in an animal or human.

The term “therapeutic effect” is art-recognized and refers to abeneficial local or systemic effect in animals, particularly mammals,and more particularly humans, caused by a pharmacologically activesubstance. The phrase “therapeutically-effective amount” means thatamount of such a substance that produces some desired local or systemiceffect at a reasonable benefit/risk ratio applicable to any treatment.The therapeutically effective amount of such substance will varydepending upon the subject and disease condition being treated, theweight and age of the subject, the severity of the disease condition,the manner of administration and the like, which can readily bedetermined by one of skill in the art. For example, certain compositionsdescribed herein may be administered in a sufficient amount to produce adesired effect at a reasonable benefit/risk ratio applicable to suchtreatment.

“Treating” a condition or disease refers to curing as well asameliorating at least one symptom of the condition or disease.

The term “vision impairment” refers to diminished vision, which is oftenonly partially reversible or irreversible upon treatment (e.g.,surgery). Particularly severe vision impairment is termed “blindness” or“vision loss”, which refers to a complete loss of vision, vision worsethan 20/200 that cannot be improved with corrective lenses, or a visualfield of less than 20 degrees diameter (10 degrees radius).

Abbreviations and Symbols

In describing the present invention, chemical elements are identified inaccordance with the Periodic Table of the Elements. Abbreviations andsymbols utilized herein are in accordance with the common usage of suchabbreviations and symbols by those skilled in the chemical andbiological arts.

Specifically, the following abbreviations may be used in the examplesand throughout the specification:

-   g (grams);-   mg (milligrams);-   kg (kilograms);-   μg (micrograms);-   L (liters);-   mL (milliliters);-   μL (microliters);-   psi (pounds per square inch);-   M (molar);-   mM (millimolar);-   μM (micromolar);-   nM (nanomolar);-   pM (picomolar);-   nm (nanometers);-   mm (millimeters);-   wt (weight);-   N (Normal);-   CFU (colony forming units);-   I. V. (intravenous);-   Hz (Hertz);-   MHz (megahertz);-   mol (moles);-   mmol (millimoles);-   RT (room temperature);-   min (minutes);-   h (hours);-   b.p. (boiling point);-   TLC (thin layer chromatography);-   T_(r) (retention time);-   RP (reverse phase);-   MeOH (methanol);-   i-PrOH (isopropanol);-   TEA (triethylamine);-   TFA (trifluoroacetic acid);-   TFAA (trifluoroacetic anhydride);-   THF (tetrahydrofuran);-   DMSO (dimethylsulfoxide);-   EtOAc (ethyl acetate);-   DME (1,2-dimethoxyethane);-   DCM (dichloromethane);-   DCE (dichloroethane);-   DMF (N,N-dimethylformamide);-   DMPU (N,N′-dimethylpropyleneurea);-   CDI (1,1-carbonyldiimidazole);-   IBCF (isobutyl chloroformate);-   AcOH (acetic acid);-   HOAt (1-hydroxy-7-azabenzotriazole);-   THP (tetrahydropyran);-   NMM (N-methylmorpholine);-   Pd/C (Palladium on Carbon);-   MTBE (tert-butyl methyl ether);-   HOBT (1-hydroxybenzotriazole);-   mCPBA (meta-chloroperbenzoic acid;-   EDC (1-[3-dimethylamino) propyl]-3-ethylcarbodiimide hydrochloride);-   Boc (tert-butyloxycarbonyl);-   FMOC (9-fluorenylmethoxycarbonyl);-   DCC (dicyclohexylcarbodiimide);-   CBZ (benzyloxycarbonyl);-   Ac (acetyl);-   atm (atmosphere);-   TMSE (2-(trimethylsilyl)ethyl);-   TMS (trimethylsilyl);-   TIPS (triisopropylsilyl);-   TB S (t-butyldimethylsilyl);-   DMAP (4-dimethylaminopyridine);-   BSA (bovine serum albumin)-   NAD (nicotinamide adenine dinucleotide);-   HPLC (high pressure liquid chromatography);-   LC/MS (liquid chromatography/mass spectrometry);-   BOP (bis(2-oxo-3-oxazolidinyl)phosphinic chloride);-   TBAF (tetra-n-butylammonium fluoride);-   HBTU (O-Benzotriazole-1-yl-N,N,N′,N′-tetramethyluroniumhexafluoro    phosphate).-   HEPES (4-(2-hydroxyethyl)-1-piperazine ethane sulfonic acid);-   DPPA (diphenylphosphoryl azide);-   LAH (Lithium aluminum hydride);-   fHNO₃ (fuming HNO₃);-   NaOMe (sodium methoxide);-   EDTA (ethylenediaminetetraacetic acid);-   TMEDA (N,N,N′,N′-tetramethyl-1,2-ethanediamine);-   NBS (N-bromosuccinimide);-   DIPEA (diisopropylethylamine);-   dppf (1,1′-bis(diphenylphosphino)ferrocene); and-   NIS (N-iodsuccinimide).

All references to ether are to diethyl ether and brine refers to asaturated aqueous solution of NaCl.

Synthetic Schemes and General Methods of Preparation

The present invention also relates to processes for making compounds ofFormulas (I) to (IV), corresponding analogs (i.e., with hydrogensubstitution at the R² position), and/or intermediate compounds thereof,respectively.

The compounds of Formulas (I) to (IV), corresponding analogs (i.e., withhydrogen substitution at the R² position) and/or intermediate compoundsthereof, or pharmaceutically acceptable salts thereof, may be obtainedby using synthetic procedures illustrated in the Schemes below or bydrawing on the knowledge of a skilled organic chemist.

The synthesis provided in these Schemes (I) to (VI) are applicable forproducing compounds of the invention having a variety of differentfunctional groups employing appropriate precursors, which are suitablyprotected if needed, to achieve compatibility with the reactionsoutlined herein. Subsequent deprotection, where needed, affordscompounds of the nature generally disclosed. While the Schemes are shownwith compounds, they are illustrative of processes that may be used tomake the compounds of the invention.

Intermediates (compounds used in the preparation of the compounds of theinvention) may also be present as salts. Thus, in reference tointermediates, the phrase “compound(s) of formula (number)” means acompound having that structural formula or a pharmaceutically acceptablesalt thereof.

The present invention also relates to processes for making compounds ofFormulas (I) to (IV), corresponding analogs (i.e., with hydrogensubstitution at the R² position), and/or intermediate compounds thereof,respectively, or pharmaceutically acceptable salts thereof.

The compounds according to Formulas (I) to (II), respectively, Thepresent invention also relates to processes for making compounds ofFormulas (I) to (IV), corresponding analogs (i.e., with hydrogensubstitution at the R² position), and/or intermediate compounds thereof,respectively, or pharmaceutically acceptable salts thereof are preparedusing conventional organic syntheses.

The compounds of the present invention may be obtained by usingsynthetic procedures illustrated in Schemes below or by drawing on theknowledge of a skilled organic chemist.

Suitable synthetic routes are depicted below in the following generalreaction schemes.

Compound Preparation

According to another embodiment, the present invention provides methodsof producing the above-defined compounds. The compounds may besynthesized using conventional techniques. Advantageously, thesecompounds are conveniently synthesized from readily available startingmaterials.

Synthetic chemistry transformations and methodologies useful insynthesizing the compounds described herein are known in the art andinclude, for example, those described in R. Larock, ComprehensiveOrganic Transformations (1989); T. W. Greene and P. G. M. Wuts,Protective Groups in Organic Synthesis, 2d. Ed. (1991); L. Fieser and M.Fieser, Fieser and Fieser's Reagents for Organic Synthesis (1994); andL. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis (1995).

General Procedures

Reagents: (a) THF, NaHCO₃, 60° C.; (b) Fe, i-PrOH, HOAc, 80° C.; (c)AlCl₃, LiAlH₄, THF, RT; (d) POCl₃, DEA, DCM, 0° C.

The commercial chloropyridine (I-1) was reacted with a nucleophilicamine (I-2) in the presence of a base (to scavenge HCl) in an aproticsolvent (eg., THF, DMF, dioxane) to provide the regioselective additionproduct (I-3). The nitro functionality of species (I-3) was reducedusing Fe(0), (see, Bechamp reduction, Org React. 2, 428, 1944) in thepresence of a Bronstead acid (HCl, HOAc) and a protic solvent. Othermetals may be used such as Sn to effect this reduction. The resultingintermediate amine species formed in situ reacted with the esterfunctionality under elevated temperatures to form the cyclic amide I-4.A strong hydride reducing agent, such as LiAlH₄, was reacted withcompound 1-4 resulting in the reduction of the ester to thecorresponding alcohol and simultaneous reduction of the lactam to acyclic amine. Reductions of this type are well-known to those instructedin the art, see H. C. Brown and S. Krishnamurthy, Tetrahedron, 1979, 35,567. Reaction of the alcohol (I-5) with an activating group (such asPOCl₃), capable of forming facile leaving group, provided the bicyclicamine compound (I-6).

Reagents: (a) TEA, CH₃OH, 120° C., 300 psi CO, PdCl₂(dppf); (b) NaH,THF, 3-(pyridin-2-yl)-2H-pyrido[1,2a][1,3,5]triazine-2,4(3H)-dione, 65°C., then H₂O; (c) DIPEA, DMF, HATU, alkyl amine, RT.

Aryl chloride (I-6) was reacted with CO under pressure and elevatedtemperature in the presence of an alcohol to produce the ester (II-1).Carbonylation reactions are described in the literature (see, Principlesand Applications of Organotransition Metal Chemistry. Sausalito, Calif.:University Science Books; 1987) and are well known to those skilled inthe art. Amine (II-1) was reacted with an acylating reagent, such astriphosgene or carbonyl diimidazole in an aprotic solvent (DCM, CHCl₃,THF, etc.) followed by treatment in situ with an aniline compound oralkyl amine in the presence of a tertiary alkyl amine base. Concommitanthydrolysis of the ester functionality occurred via in situ generatedNaOH to form the urea species (II-2). Carboxylic acid (II-2) was reactedwith an alkyl amine in the presense of a coupling reagent (HATU) in apolar aprotic solvent to give the corresponding amide (II-3). A varietyof amide coupling reagents such as EDC, PyBrop, etc. are commerciallyavailable. Amide coupling reactions are generally run in solvents suchas DCM or DMF, utilizing an organic base like Et₃N or (i-Pr)₂NEt.

Reagents: (a) Pd₂(dba)₃, X-Phos, Cs₂CO₃, dioxane/H₂O, 90° C.; (b) TEA,triphosgene, THF; then 2-aminopyridine, 65° C.

The chloro functionality of compound 1-6 was coupled with a boronic acidusing Suzuki coupling chemistry to give III-1. Suzuki-like couplings aretypically run using a palladium(0) catalyst such as Pd(PPh₃)₄ with aninorganic base, for example K₂CO₃, Na₂CO₃ or K₃PO₄, in an aqueousmixture containing ethereal solvents such as DME, dioxane, or THF.Methods for palladium-mediated couplings are described in standardreference volumes, such as Schlosser “Organometallics in Synthesis”(published by Wiley and sons). Compound III-1 was reacted with anacylating reagent, such as triphosgene or carbonyl diimidazole in anaprotic solvent (DCM, CHCl₃, THF, etc.) to give a reactive acylintermediate species which was treated in situ with an aniline compoundor alkyl amine in the presence of a tertiary alkyl amine base to formthe urea species (III-2).

Reagents: (a) 3-trifluoromethylpyrrolidine,(1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene)chloro)(3-phenylallyl)palladium, KOt-Bu, DME, 80° C.; (b) TEA, triphosgene,THF; then 2-aminopyridine, 65° C.

The chloro functionality of urea 1-6 was selectively displaced by analkyl amine using Buchwald-Hartwig amination conditions to give (IV-1).Buchwald-Hartwig reactions are typically run using a palladium(0)catalyst such as Pd(PPh₃)₄ with an bulky Bronstead base, for exampleKOt-Bu or KHMDS, containing ethereal solvents such as DME, dioxane, orTHF. Methods for palladium-mediated amine couplings are described inHartwig, J. F. (1998), Transition Metal Catalyzed Synthesis ofArylamines and Aryl Ethers from Aryl Halides and Triflates: Scope andMechanism, Angew. Chem. Int. Ed. 37: 2046-2067. Compound IV-1 wasreacted with an acylating reagent, such as triphosgene or carbonyldiimidazole in an aprotic solvent (DCM, CHCl₃, THF, etc.) to give areactive acyl intermediate species which was treated in situ with ananiline compound in the presence of a tertiary alkyl amine base to formthe urea species (IV-2).

Reagents: (a) NBS or NCS, CHCl₃, 60° C.; (b) NaH, THF,3-(pyridin-2-yl)-2H-pyrido[1,2a][1,3,5]triazine-2,4(3H)-dione, 65° C.,then H₂O; (c) DIPEA, DMF, HATU, alkyl amine, RT.

Amine (II-1) was reacted with an electrophilic halogenating reagent,such as NCS or NBS, in an appropriate solvent to give the correspondinghalogenated species V-1 in a regioselective manner. Many methods existto effect the halogenation of an aromatic ring and are well-known tothose skilled in the art. Ester (V-1) was reacted with an acylatingreagent, such as triphosgene or carbonyl diimidazole in an aproticsolvent (DCM, CHCl₃, THF, etc.) followed by treatment with an anilinecompound or alkyl amine in the presence of a tertiary alkyl amine base.Concommitant hydrolysis of the ester functionality occurred via in situgenerated NaOH to form the urea species (V-2). Carboxylic acid (V-2) wasreacted with an alkyl amine in the presense of a coupling reagent (HATU)in a polar aprotic solvent to give the corresponding amide (V-3). Avariety of amide coupling reagents such as EDC, PyBrop, etc. arecommercially available. Amide coupling reactions are generally run insolvents such as DCM or DMF, utilizing an organic base like Et₃N or(i-Pr)₂NEt.

Reagents: (a) NBS or NCS, CHCl₃, 60° C.; (b) TEA, triphosgene, THF; then2-aminopyridine, 65° C.

Compound III-1 was reacted with an electrophilic halogenating reagent,such as NCS or NBS, in an appropriate solvent to give the correspondinghalogenated species VI-1 in a regioselective manner. Many methods existto effect the halogenation of an aromatic ring and are well-known tothose skilled in the art. Compound VI-1 was reacted with an acylatingreagent, such as triphosgene or carbonyl diimidazole in an aproticsolvent (DCM, CHCl₃, THF, etc.) to give a reactive acyl intermediatespecies which was treated in situ with an aniline compound or alkylamine in the presence of a tertiary alkyl amine base to form the ureaspecies (VI-2).

Reagents: (a) NIS, CHCl₃, 60° C.; (b) trimethylboroxine, Pd(PPh₃)₄,K₂CO₃, dioxane/H₂O, 90° C.;

Compound 1-6 was reacted with an electrophilic halogenating reagent(NIS) in an appropriate solvent to give the corresponding halogenatedspecies VII-1 in a regioselective manner. Many methods exist to effectthe halogenation of an aromatic ring and are well-known to those skilledin the art. The iodo functionality of compound VII-1 was coupled with analkylboronic acid using Suzuki coupling chemistry to give VII-2.Suzuki-like couplings are typically run using a palladium(0) catalystsuch as Pd(PPh₃)₄ with an inorganic base, for example K₂CO₃, Na₂CO₃ orK₃PO₄, in an aqueous mixture containing ethereal solvents such as DME,dioxane, or THF. Methods for palladium-mediated couplings are describedin standard reference volumes, such as Schlosser “Organometallics inSynthesis” (published by Wiley and sons). The methylated species (VII-2)was used in schemes II and III as a surrogate for compound 1-6 toproduce the analogous methylated products.

Compound Characteristics and Properties

In an exemplary embodiment, a therapeutic compound may traverse thecytoplasmic membrane of a cell. For example, a compound may have acell-permeability of at least about 20%, 50%, 75%, 80%, 90% or 95%.

Compounds described herein may also have one or more of the followingcharacteristics: the compound may be essentially non-toxic to a cell orsubject; the compound may be an organic molecule or a small molecule of2000 amu or less, 1000 amu or less; a compound may have a half-lifeunder normal atmospheric conditions of at least about 30 days, 60 days,120 days, 6 months or 1 year; the compound may have a half-life insolution of at least about 30 days, 60 days, 120 days, 6 months or 1year; a compound may be more stable in solution than resveratrol by atleast a factor of about 50%, 2 fold, 5 fold, 10 fold, 30 fold, 50 foldor 100 fold; a compound may promote deacetylation of the DNA repairfactor Ku70; a compound may promote deacetylation of RelA/p65; acompound may increase general turnover rates and enhance the sensitivityof cells to TNF-induced apoptosis.

In certain embodiments, a sirtuin-modulating compound does not have anysubstantial ability to inhibit a histone deacetylase (HDAC) class I,and/or an HDAC class II at concentrations (e.g., in vivo) effective formodulating the deacetylase activity of the sirtuin. For instance, inpreferred embodiments, the sirtuin-modulating compound is asirtuin-modulating compound and is chosen to have an EC₅₀ for activatingsirtuin deacetylase activity that is at least 5 fold less than the EC₅₀for inhibition of an HDAC I and/or HDAC II, and even more preferably atleast 10 fold, 100 fold or even 1000 fold less. Methods for assayingHDAC I and/or HDAC II activity are well known in the art and kits toperform such assays may be purchased commercially. See e.g., BioVision,Inc. (Mountain View, Calif.; world wide web at biovision.com) and ThomasScientific (Swedesboro, N.J.; world wide web at tomassci.com).

In certain embodiments, a sirtuin-modulating compound does not have anysubstantial ability to modulate sirtuin homologs. In certainembodiments, an activator of a human sirtuin protein may not have anysubstantial ability to activate a sirtuin protein from lower eukaryotes,particularly yeast or human pathogens, at concentrations (e.g., in vivo)effective for activating the deacetylase activity of human sirtuin. Forexample, a sirtuin-modulating compound may be chosen to have an EC₅₀ foractivating a human sirtuin, such as SIRT1 and/or SIRT3, deacetylaseactivity that is at least 5 fold less than the EC₅₀ for activating ayeast sirtuin, such as Sir2 (such as Candida, S. cerevisiae, etc.), andeven more preferably at least 10 fold, 100 fold or even 1000 fold less.In another embodiment, an inhibitor of a sirtuin protein from lowereukaryotes, particularly yeast or human pathogens, does not have anysubstantial ability to inhibit a sirtuin protein from humans atconcentrations (e.g., in vivo) effective for inhibiting the deacetylaseactivity of a sirtuin protein from a lower eukaryote. For example, asirtuin-inhibiting compound may be chosen to have an IC₅₀ for inhibitinga human sirtuin, such as SIRT1 and/or SIRT3, deacetylase activity thatis at least 5 fold less than the IC₅₀ for inhibiting a yeast sirtuin,such as Sir2 (such as Candida, S. cerevisiae, etc.), and even morepreferably at least 10 fold, 100 fold or even 1000 fold less.

In certain embodiments, a sirtuin-modulating compound may have theability to modulate one or more sirtuin protein homologs, such as, forexample, one or more of human SIRT1, SIRT2, SIRT3, SIRT4, SIRT5, SIRT6,or SIRT7. In some embodiments, a sirtuin-modulating compound has theability to modulate both a SIRT1 and a SIRT3 protein.

In other embodiments, a SIRT1 modulator does not have any substantialability to modulate other sirtuin protein homologs, such as, forexample, one or more of human SIRT2, SIRT3, SIRT4, SIRT5, SIRT6, orSIRT7, at concentrations (e.g., in vivo) effective for modulating thedeacetylase activity of human SIRT1. For example, a sirtuin-modulatingcompound may be chosen to have an ED₅₀ for modulating human SIRT1deacetylase activity that is at least 5 fold less than the ED₅₀ formodulating one or more of human SIRT2, SIRT3, SIRT4, SIRT5, SIRT6, orSIRT7, and even more preferably at least 10 fold, 100 fold or even 1000fold less. In some embodiments, a SIRT1 modulator does not have anysubstantial ability to modulate a SIRT3 protein.

In other embodiments, a SIRT3 modulator does not have any substantialability to modulate other sirtuin protein homologs, such as, forexample, one or more of human SIRT1, SIRT2, SIRT4, SIRT5, SIRT6, orSIRT7, at concentrations (e.g., in vivo) effective for modulating thedeacetylase activity of human SIRT3. For example, a sirtuin-modulatingcompound may be chosen to have an ED₅₀ for modulating human SIRT3deacetylase activity that is at least 5 fold less than the ED₅₀ formodulating one or more of human SIRT1, SIRT2, SIRT4, SIRT5, SIRT6, orSIRT7, and even more preferably at least 10 fold, 100 fold or even 1000fold less. In some embodiments, a SIRT3 modulator does not have anysubstantial ability to modulate a SIRT1 protein.

In certain embodiments, a sirtuin-modulating compound may have a bindingaffinity for a sirtuin protein of about 10⁻⁹M, 10⁻¹⁰M, 10⁻¹¹M, 10⁻¹²M orless. A sirtuin-modulating compound may reduce (activator) or increase(inhibitor) the apparent Km of a sirtuin protein for its substrate orNAD⁺ (or other cofactor) by a factor of at least about 2, 3, 4, 5, 10,20, 30, 50 or 100. In certain embodiments, Km values are determinedusing the mass spectrometry assay described herein. Preferred activatingcompounds reduce the Km of a sirtuin for its substrate or cofactor to agreater extent than caused by resveratrol at a similar concentration orreduce the Km of a sirtuin for its substrate or cofactor similar to thatcaused by resveratrol at a lower concentration. A sirtuin-modulatingcompound may increase the Vmax of a sirtuin protein by a factor of atleast about 2, 3, 4, 5, 10, 20, 30, 50 or 100. A sirtuin-modulatingcompound may have an ED₅₀ for modulating the deacetylase activity of aSIRT1 and/or SIRT3 protein of less than about 1 nM, less than about 10nM, less than about 100 nM, less than about 1 μM, less than about 10 μM,less than about 100 μM, or from about 1-10 nM, from about 10-100 nM,from about 0.1-1 μM, from about 1-10 μM or from about 10-100 μM. Asirtuin-modulating compound may modulate the deacetylase activity of aSIRT1 and/or SIRT3 protein by a factor of at least about 5, 10, 20, 30,50, or 100, as measured in a cellular assay or in a cell based assay. Asirtuin-modulating compound may cause at least about 10%, 30%, 50%, 80%,2 fold, 5 fold, 10 fold, 50 fold or 100 fold greater induction of thedeacetylase activity of a sirtuin protein relative to the sameconcentration of resveratrol. A sirtuin-modulating compound may have anED₅₀ for modulating SIRT5 that is at least about 10 fold, 20 fold, 30fold, 50 fold greater than that for modulating SIRT1 and/or SIRT3.

Exemplary Uses

In certain aspects, the invention provides methods or uses formodulating the level and/or activity of a sirtuin protein and methods ofuse thereof.

In certain embodiments, the invention provides methods or uses for usingsirtuin-modulating compounds wherein the sirtuin-modulating compoundsactivate a sirtuin protein, e.g., increase the level and/or activity ofa sirtuin protein. Sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be useful for a variety oftherapeutic applications including, for example, increasing the lifespanof a cell, and treating and/or preventing a wide variety of diseases anddisorders including, for example, diseases or disorders related to agingor stress, diabetes, obesity, neurodegenerative diseases, cardiovasculardisease, blood clotting disorders, inflammation, cancer, and/orflushing, etc. The methods or uses comprise administering to a subjectin need thereof a pharmaceutically effective amount of asirtuin-modulating compound, e.g., a sirtuin-modulating compound.

Without wishing to be bound by theory, it is believed that activators ofthe instant invention may interact with a sirtuin at the same locationwithin the sirtuin protein (e.g., active site or site affecting the Kmor Vmax of the active site). It is believed that this is the reason whycertain classes of sirtuin activators and inhibitors can havesubstantial structural similarity.

In certain embodiments, the sirtuin-modulating compounds describedherein may be taken alone or in combination with other compounds. Incertain embodiments, a mixture of two or more sirtuin-modulatingcompounds may be administered to a subject in need thereof.

In another embodiment, a sirtuin-modulating compound that increases thelevel and/or activity of a sirtuin protein may be administered with oneor more of the following compounds: resveratrol, butein, fisetin,piceatannol, or quercetin.

In an exemplary embodiment, a sirtuin-modulating compound that increasesthe level and/or activity of a sirtuin protein may be administered incombination with nicotinic acid or nicotinamide riboside. In anotherembodiment, a sirtuin-modulating compound that decreases the leveland/or activity of a sirtuin protein may be administered with one ormore of the following compounds: nicotinamide (NAM), suramin; NF023 (aG-protein antagonist); NF279 (a purinergic receptor antagonist); Trolox(6-hydroxy-2,5,7,8,tetramethylchroman-2-carboxylic acid);(−)-epigallocatechin (hydroxy on sites 3,5,7,3′,4′,5′);(−)-epigallocatechin gallate (Hydroxy sites 5,7,3′,4′,5′ and gallateester on 3); cyanidin chloride (3,5,7,3′,4′-pentahydroxyflavyliumchloride); delphinidin chloride (3,5,7,3′,4′,5′-hexahydroxyflavyliumchloride); myricetin (cannabiscetin; 3,5,7,3′,4′,5′-hexahydroxyflavone);3,7,3′,4′,5′-pentahydroxyflavone; gossypetin(3,5,7,8,3′,4′-hexahydroxyflavone), sirtinol; and splitomicin.

In yet another embodiment, one or more sirtuin-modulating compounds maybe administered with one or more therapeutic agents for the treatment orprevention of various diseases, including, for example, cancer,diabetes, neurodegenerative diseases, cardiovascular disease, bloodclotting, inflammation, flushing, obesity, aging, stress, etc. Invarious embodiments, combination therapies comprising asirtuin-modulating compound may refer to (1) pharmaceutical compositionsthat comprise one or more sirtuin-modulating compounds in combinationwith one or more therapeutic agents (e.g., one or more therapeuticagents described herein); and (2) co-administration of one or moresirtuin-modulating compounds with one or more therapeutic agents whereinthe sirtuin-modulating compound and therapeutic agent have not beenformulated in the same compositions (but may be present within the samekit or package, such as a blister pack or other multi-chamber package;connected, separately sealed containers (e.g., foil pouches) that can beseparated by the user; or a kit where the compound(s) and othertherapeutic agent(s) are in separate vessels). When using separateformulations, the sirtuin-modulating compound may be administeredsimultaneous with, intermittent with, staggered with, prior to,subsequent to, or combinations thereof, the administration of anothertherapeutic agent.

In certain embodiments, methods or uses for reducing, preventing ortreating diseases or disorders using a compound described herein mayalso comprise increasing the protein level of a sirtuin, such as humanSIRT1, SIRT2 and/or SIRT3, or homologs thereof. Increasing proteinlevels can be achieved by introducing into a cell one or more copies ofa nucleic acid that encodes a sirtuin. For example, the level of asirtuin can be increased in a mammalian cell by introducing into themammalian cell a nucleic acid encoding the sirtuin, e.g., increasing thelevel of SIRT1 by introducing a nucleic acid encoding the amino acidsequence set forth in GenBank Accession No. NP_036370 and/or increasingthe level of SIRT3 by introducing a nucleic acid encoding the amino acidsequence set forth in GenBank Accession No. AAH01042.

A nucleic acid that is introduced into a cell to increase the proteinlevel of a sirtuin may encode a protein that is at least about 80%, 85%,90%, 95%, 98%, or 99% identical to the sequence of a sirtuin, e.g.,SIRT1 and/or SIRT3 protein. For example, the nucleic acid encoding theprotein may be at least about 80%, 85%, 90%, 95%, 98%, or 99% identicalto a nucleic acid encoding a SIRT1 (e.g. GenBank Accession No.NM_012238) and/or SIRT3 (e.g., GenBank Accession No. BC001042) protein.The nucleic acid may also be a nucleic acid that hybridizes, preferablyunder stringent hybridization conditions, to a nucleic acid encoding awild-type sirtuin, e.g., SIRT1 and/or SIRT3 protein. Stringenthybridization conditions may include hybridization and a wash in 0.2×SSCat 65° C. When using a nucleic acid that encodes a protein that isdifferent from a wild-type sirtuin protein, such as a protein that is afragment of a wild-type sirtuin, the protein is preferably biologicallyactive, e.g., is capable of deacetylation. It is only necessary toexpress in a cell a portion of the sirtuin that is biologically active.For example, a protein that differs from wild-type SIRT1 having GenBankAccession No. NP_036370, preferably contains the core structure thereof.The core structure sometimes refers to amino acids 62-293 of GenBankAccession No. NP_036370, which are encoded by nucleotides 237 to 932 ofGenBank Accession No. NM_012238, which encompasses the NAD binding aswell as the substrate binding domains. The core domain of SIRT1 may alsorefer to about amino acids 261 to 447 of GenBank Accession No.NP_036370, which are encoded by nucleotides 834 to 1394 of GenBankAccession No. NM_012238; to about amino acids 242 to 493 of GenBankAccession No. NP_036370, which are encoded by nucleotides 777 to 1532 ofGenBank Accession No. NM_012238; or to about amino acids 254 to 495 ofGenBank Accession No. NP_036370, which are encoded by nucleotides 813 to1538 of GenBank Accession No. NM_012238. Whether a protein retains abiological function, e.g., deacetylation capabilities, can be determinedaccording to methods known in the art.

In certain embodiments, methods or uses for reducing, preventing ortreating diseases or disorders using a sirtuin-modulating compound mayalso comprise decreasing the protein level of a sirtuin, such as humanSIRT1, SIRT2 and/or SIRT3, or homologs thereof. Decreasing a sirtuinprotein level can be achieved according to methods known in the art. Forexample, an siRNA, an antisense nucleic acid, or a ribozyme targeted tothe sirtuin can be expressed in the cell. A dominant negative sirtuinmutant, e.g., a mutant that is not capable of deacetylating, may also beused. For example, mutant H363Y of SIRT1, described, e.g., in Luo et al.(2001) Cell 107:137 can be used. Alternatively, agents that inhibittranscription can be used.

Methods or uses for modulating sirtuin protein levels also includemethods or uses for modulating the transcription of genes encodingsirtuins, methods for stabilizing/destabilizing the corresponding mRNAs,and other methods known in the art.

Aging/Stress

In one aspect, the invention provides a method extending the lifespan ofa cell, extending the proliferative capacity of a cell, slowing aging ofa cell, promoting the survival of a cell, delaying cellular senescencein a cell, mimicking the effects of calorie restriction, increasing theresistance of a cell to stress, or preventing apoptosis of a cell, bycontacting the cell with a sirtuin-modulating compound of the inventionthat increases the level and/or activity of a sirtuin protein. In anexemplary embodiment, the methods or uses comprise contacting the cellwith a sirtuin-modulating compound.

The methods or uses described herein may be used to increase the amountof time that cells, particularly primary cells (i.e., cells obtainedfrom an organism, e.g., a human), may be kept alive in a cell culture.Embryonic stem (ES) cells and pluripotent cells, and cellsdifferentiated therefrom, may also be treated with a sirtuin-modulatingcompound that increases the level and/or activity of a sirtuin proteinto keep the cells, or progeny thereof, in culture for longer periods oftime. Such cells can also be used for transplantation into a subject,e.g., after ex vivo modification.

In one aspect, cells that are intended to be preserved for long periodsof time may be treated with a sirtuin-modulating compound that increasesthe level and/or activity of a sirtuin protein. The cells may be insuspension (e.g., blood cells, serum, biological growth media, etc.) orin tissues or organs. For example, blood collected from an individualfor purposes of transfusion may be treated with a sirtuin-modulatingcompound that increases the level and/or activity of a sirtuin proteinto preserve the blood cells for longer periods of time. Additionally,blood to be used for forensic purposes may also be preserved using asirtuin-modulating compound that increases the level and/or activity ofa sirtuin protein. Other cells that may be treated to extend theirlifespan or protect against apoptosis include cells for consumption,e.g., cells from non-human mammals (such as meat) or plant cells (suchas vegetables).

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may also be applied during developmental and growthphases in mammals, plants, insects or microorganisms, in order to, e.g.,alter, retard or accelerate the developmental and/or growth process.

In another aspect, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be used to treat cells usefulfor transplantation or cell therapy, including, for example, solidtissue grafts, organ transplants, cell suspensions, stem cells, bonemarrow cells, etc. The cells or tissue may be an autograft, anallograft, a syngraft or a xenograft. The cells or tissue may be treatedwith the sirtuin-modulating compound prior toadministration/implantation, concurrently withadministration/implantation, and/or post administration/implantationinto a subject. The cells or tissue may be treated prior to removal ofthe cells from the donor individual, ex vivo after removal of the cellsor tissue from the donor individual, or post implantation into therecipient. For example, the donor or recipient individual may be treatedsystemically with a sirtuin-modulating compound or may have a subset ofcells/tissue treated locally with a sirtuin-modulating compound thatincreases the level and/or activity of a sirtuin protein. In certainembodiments, the cells or tissue (or donor/recipient individuals) mayadditionally be treated with another therapeutic agent useful forprolonging graft survival, such as, for example, an immunosuppressiveagent, a cytokine, an angiogenic factor, etc.

In yet other embodiments, cells may be treated with a sirtuin-modulatingcompound that increases the level and/or activity of a sirtuin proteinin vivo, e.g., to increase their lifespan or prevent apoptosis. Forexample, skin can be protected from aging (e.g., developing wrinkles,loss of elasticity, etc.) by treating skin or epithelial cells with asirtuin-modulating compound that increases the level and/or activity ofa sirtuin protein. In an exemplary embodiment, skin is contacted with apharmaceutical or cosmetic composition comprising a sirtuin-modulatingcompound that increases the level and/or activity of a sirtuin protein.Exemplary skin afflictions or skin conditions that may be treated inaccordance with the methods or uses described herein include disordersor diseases associated with or caused by inflammation, sun damage ornatural aging. For example, the compositions find utility in theprevention or treatment of contact dermatitis (including irritantcontact dermatitis and allergic contact dermatitis), atopic dermatitis(also known as allergic eczema), actinic keratosis, keratinizationdisorders (including eczema), epidermolysis bullosa diseases (includingpemphigus), exfoliative dermatitis, seborrheic dermatitis, erythemas(including erythema multiforme and erythema nodosum), damage caused bythe sun or other light sources, discoid lupus erythematosus,dermatomyositis, psoriasis, skin cancer and the effects of naturalaging. In another embodiment, sirtuin-modulating compounds that increasethe level and/or activity of a sirtuin protein may be used for thetreatment of wounds and/or burns to promote healing, including, forexample, first-, second- or third-degree burns and/or thermal, chemicalor electrical burns. The formulations may be administered topically, tothe skin or mucosal tissue.

Topical formulations comprising one or more sirtuin-modulating compoundsthat increase the level and/or activity of a sirtuin protein may also beused as preventive, e.g., chemopreventive, compositions. When used in achemopreventive method, susceptible skin is treated prior to any visiblecondition in a particular individual.

Sirtuin-modulating compounds may be delivered locally or systemically toa subject. In certain embodiments, a sirtuin-modulating compound isdelivered locally to a tissue or organ of a subject by injection,topical formulation, etc.

In another embodiment, a sirtuin-modulating compound that increases thelevel and/or activity of a sirtuin protein may be used for treating orpreventing a disease or condition induced or exacerbated by cellularsenescence in a subject; methods or uses for decreasing the rate ofsenescence of a subject, e.g., after onset of senescence; methods forextending the lifespan of a subject; methods or uses for treating orpreventing a disease or condition relating to lifespan; methods or usesfor treating or preventing a disease or condition relating to theproliferative capacity of cells; and methods or uses for treating orpreventing a disease or condition resulting from cell damage or death.In certain embodiments, the method does not act by decreasing the rateof occurrence of diseases that shorten the lifespan of a subject. Incertain embodiments, a method does not act by reducing the lethalitycaused by a disease, such as cancer.

In yet another embodiment, a sirtuin-modulating compound that increasesthe level and/or activity of a sirtuin protein may be administered to asubject in order to generally increase the lifespan of its cells and toprotect its cells against stress and/or against apoptosis. It isbelieved that treating a subject with a compound described herein issimilar to subjecting the subject to hormesis, i.e., mild stress that isbeneficial to organisms and may extend their lifespan.

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may be administered to a subject to prevent aging andaging-related consequences or diseases, such as stroke, heart disease,heart failure, arthritis, high blood pressure, and Alzheimer's disease.Other conditions that can be treated include ocular disorders, e.g.,associated with the aging of the eye, such as cataracts, glaucoma, andmacular degeneration. Sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein can also be administered tosubjects for treatment of diseases, e.g., chronic diseases, associatedwith cell death, in order to protect the cells from cell death.Exemplary diseases include those associated with neural cell death,neuronal dysfunction, or muscular cell death or dysfunction, such asParkinson's disease, Alzheimer's disease, multiple sclerosis,amyotrophic lateral sclerosis, and muscular dystrophy; AIDS; fulminanthepatitis; diseases linked to degeneration of the brain, such asCreutzfeld-Jakob disease, retinitis pigmentosa and cerebellardegeneration; myelodysplasia such as aplastic anemia; ischemic diseasessuch as myocardial infarction and stroke; hepatic diseases such asalcoholic hepatitis, hepatitis B and hepatitis C; joint-diseases such asosteoarthritis; atherosclerosis; alopecia; damage to the skin due to UVlight; lichen planus; atrophy of the skin; cataract; and graftrejections. Cell death can also be caused by surgery, drug therapy,chemical exposure or radiation exposure.

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein can also be administered to a subject suffering froman acute disease, e.g., damage to an organ or tissue, e.g., a subjectsuffering from stroke or myocardial infarction or a subject sufferingfrom a spinal cord injury. Sirtuin-modulating compounds that increasethe level and/or activity of a sirtuin protein may also be used torepair an alcoholic's liver.

Cardiovascular Disease

In another embodiment, the invention provides a method for treatingand/or preventing a cardiovascular disease by administering to a subjectin need thereof a sirtuin-modulating compound that increases the leveland/or activity of a sirtuin protein.

Cardiovascular diseases that can be treated or prevented using thesirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein include cardiomyopathy or myocarditis; such asidiopathic cardiomyopathy, metabolic cardiomyopathy, alcoholiccardiomyopathy, drug-induced cardiomyopathy, ischemic cardiomyopathy,and hypertensive cardiomyopathy. Also treatable or preventable usingcompounds and methods or uses described herein are atheromatousdisorders of the major blood vessels (macrovascular disease) such as theaorta, the coronary arteries, the carotid arteries, the cerebrovasculararteries, the renal arteries, the iliac arteries, the femoral arteries,and the popliteal arteries. Other vascular diseases that can be treatedor prevented include those related to platelet aggregation, the retinalarterioles, the glomerular arterioles, the vasa nervorum, cardiacarterioles, and associated capillary beds of the eye, the kidney, theheart, and the central and peripheral nervous systems. Thesirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may also be used for increasing HDL levels in plasmaof an individual.

Yet other disorders that may be treated with sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteininclude restenosis, e.g., following coronary intervention, and disordersrelating to an abnormal level of high density and low densitycholesterol.

In certain embodiments, a sirtuin-modulating compound that increases thelevel and/or activity of a sirtuin protein may be administered as partof a combination therapy with another cardiovascular agent. In certainembodiments, a sirtuin-modulating compound that increases the leveland/or activity of a sirtuin protein may be administered as part of acombination therapy with an anti-arrhythmia agent. In anotherembodiment, a sirtuin-modulating compound that increases the leveland/or activity of a sirtuin protein may be administered as part of acombination therapy with another cardiovascular agent.

Cell Death/Cancer

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may be administered to subjects who have recentlyreceived or are likely to receive a dose of radiation or toxin. Incertain embodiments, the dose of radiation or toxin is received as partof a work-related or medical procedure, e.g., administered as aprophylactic measure. In another embodiment, the radiation or toxinexposure is received unintentionally. In such a case, the compound ispreferably administered as soon as possible after the exposure toinhibit apoptosis and the subsequent development of acute radiationsyndrome.

Sirtuin-modulating compounds may also be used for treating and/orpreventing cancer. In certain embodiments, sirtuin-modulating compoundsthat increase the level and/or activity of a sirtuin protein may be usedfor treating and/or preventing cancer. Calorie restriction has beenlinked to a reduction in the incidence of age-related disordersincluding cancer. Accordingly, an increase in the level and/or activityof a sirtuin protein may be useful for treating and/or preventing theincidence of age-related disorders, such as, for example, cancer.Exemplary cancers that may be treated using a sirtuin-modulatingcompound are those of the brain and kidney; hormone-dependent cancersincluding breast, prostate, testicular, and ovarian cancers; lymphomas,and leukemias. In cancers associated with solid tumors, a modulatingcompound may be administered directly into the tumor. Cancer of bloodcells, e.g., leukemia, can be treated by administering a modulatingcompound into the blood stream or into the bone marrow. Benign cellgrowth, e.g., warts, can also be treated. Other diseases that can betreated include autoimmune diseases, e.g., systemic lupus erythematosus,scleroderma, and arthritis, in which autoimmune cells should be removed.Viral infections such as herpes, HIV, adenovirus, and HTLV-1 associatedmalignant and benign disorders can also be treated by administration ofsirtuin-modulating compound. Alternatively, cells can be obtained from asubject, treated ex vivo to remove certain undesirable cells, e.g.,cancer cells, and administered back to the same or a different subject.

Chemotherapeutic agents may be co-administered with modulating compoundsdescribed herein as having anti-cancer activity, e.g., compounds thatinduce apoptosis, compounds that reduce lifespan or compounds thatrender cells sensitive to stress. Chemotherapeutic agents may be used bythemselves with a sirtuin-modulating compound described herein asinducing cell death or reducing lifespan or increasing sensitivity tostress and/or in combination with other chemotherapeutics agents. Inaddition to conventional chemotherapeutics, the sirtuin-modulatingcompounds described herein may also be used with antisense RNA, RNAi orother polynucleotides to inhibit the expression of the cellularcomponents that contribute to unwanted cellular proliferation.

Combination therapies comprising sirtuin-modulating compounds and aconventional chemotherapeutic agent may be advantageous over combinationtherapies known in the art because the combination allows theconventional chemotherapeutic agent to exert greater effect at lowerdosage. In a preferred embodiment, the effective dose (ED₅₀) for achemotherapeutic agent, or combination of conventional chemotherapeuticagents, when used in combination with a sirtuin-modulating compound isat least 2 fold less than the ED₅₀ for the chemotherapeutic agent alone,and even more preferably at 5 fold, 10 fold or even 25 fold less.Conversely, the therapeutic index (TI) for such chemotherapeutic agentor combination of such chemotherapeutic agent when used in combinationwith a sirtuin-modulating compound described herein can be at least 2fold greater than the TI for conventional chemotherapeutic regimenalone, and even more preferably at 5 fold, 10 fold or even 25 foldgreater.

Neuronal Diseases/Disorders

In certain aspects, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein can be used to treat patientssuffering from neurodegenerative diseases, and traumatic or mechanicalinjury to the central nervous system (CNS), spinal cord or peripheralnervous system (PNS). Neurodegenerative disease typically involvesreductions in the mass and volume of the human brain, which may be dueto the atrophy and/or death of brain cells, which are far more profoundthan those in a healthy person that are attributable to aging.Neurodegenerative diseases can evolve gradually, after a long period ofnormal brain function, due to progressive degeneration (e.g., nerve celldysfunction and death) of specific brain regions. Alternatively,neurodegenerative diseases can have a quick onset, such as thoseassociated with trauma or toxins. The actual onset of brain degenerationmay precede clinical expression by many years. Examples ofneurodegenerative diseases include, but are not limited to, Alzheimer'sdisease (AD), Parkinson's disease (PD), Huntington's disease (HD),amyotrophic lateral sclerosis (ALS; Lou Gehrig's disease), diffuse Lewybody disease, chorea-acanthocytosis, primary lateral sclerosis, oculardiseases (ocular neuritis), chemotherapy-induced neuropathies (e.g.,from vincristine, paclitaxel, bortezomib), diabetes-induced neuropathiesand Friedreich's ataxia. Sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein can be used to treat thesedisorders and others as described below.

AD is a CNS disorder that results in memory loss, unusual behavior,personality changes, and a decline in thinking abilities. These lossesare related to the death of specific types of brain cells and thebreakdown of connections and their supporting network (e.g. glial cells)between them. The earliest symptoms include loss of recent memory,faulty judgment, and changes in personality. PD is a CNS disorder thatresults in uncontrolled body movements, rigidity, tremor, anddyskinesia, and is associated with the death of brain cells in an areaof the brain that produces dopamine. ALS (motor neuron disease) is a CNSdisorder that attacks the motor neurons, components of the CNS thatconnect the brain to the skeletal muscles.

HD is another neurodegenerative disease that causes uncontrolledmovements, loss of intellectual faculties, and emotional disturbance.Tay-Sachs disease and Sandhoff disease are glycolipid storage diseaseswhere GM2 ganglioside and related glycolipids substrates forβ-hexosaminidase accumulate in the nervous system and trigger acuteneurodegeneration.

It is well-known that apoptosis plays a role in AIDS pathogenesis in theimmune system. However, HIV-1 also induces neurological disease, whichcan be treated with sirtuin-modulating compounds of the invention.

Neuronal loss is also a salient feature of prion diseases, such asCreutzfeldt-Jakob disease in human, BSE in cattle (mad cow disease),Scrapie Disease in sheep and goats, and feline spongiform encephalopathy(FSE) in cats. Sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be useful for treating orpreventing neuronal loss due to these prior diseases.

In another embodiment, a sirtuin-modulating compound that increases thelevel and/or activity of a sirtuin protein may be used to treat orprevent any disease or disorder involving axonopathy. Distal axonopathyis a type of peripheral neuropathy that results from some metabolic ortoxic derangement of peripheral nervous system (PNS) neurons. It is themost common response of nerves to metabolic or toxic disturbances, andas such may be caused by metabolic diseases such as diabetes, renalfailure, deficiency syndromes such as malnutrition and alcoholism, orthe effects of toxins or drugs. Those with distal axonopathies usuallypresent with symmetrical glove-stocking sensori-motor disturbances. Deeptendon reflexes and autonomic nervous system (ANS) functions are alsolost or diminished in affected areas.

Diabetic neuropathies are neuropathic disorders that are associated withdiabetes mellitus. Relatively common conditions which may be associatedwith diabetic neuropathy include third nerve palsy; mononeuropathy;mononeuritis multiplex; diabetic amyotrophy; a painful polyneuropathy;autonomic neuropathy; and thoracoabdominal neuropathy.

Peripheral neuropathy is the medical term for damage to nerves of theperipheral nervous system, which may be caused either by diseases of thenerve or from the side-effects of systemic illness. Major causes ofperipheral neuropathy include seizures, nutritional deficiencies, andHIV, though diabetes is the most likely cause.

In an exemplary embodiment, a sirtuin-modulating compound that increasesthe level and/or activity of a sirtuin protein may be used to treat orprevent multiple sclerosis (MS), including relapsing MS andmonosymptomatic MS, and other demyelinating conditions, such as, forexample, chronic inflammatory demyelinating polyneuropathy (CIDP), orsymptoms associated therewith.

In yet another embodiment, a sirtuin-modulating compound that increasesthe level and/or activity of a sirtuin protein may be used to treattrauma to the nerves, including, trauma due to disease, injury(including surgical intervention), or environmental trauma (e.g.,neurotoxins, alcoholism, etc.).

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may also be useful to prevent, treat, and alleviatesymptoms of various PNS disorders. The term “peripheral neuropathy”encompasses a wide range of disorders in which the nerves outside of thebrain and spinal cord—peripheral nerves—have been damaged. Peripheralneuropathy may also be referred to as peripheral neuritis, or if manynerves are involved, the terms polyneuropathy or polyneuritis may beused.

PNS diseases treatable with sirtuin-modulating compounds that increasethe level and/or activity of a sirtuin protein include: diabetes,leprosy, Charcot-Marie-Tooth disease, Guillain-Barré syndrome andBrachial Plexus Neuropathies (diseases of the cervical and firstthoracic roots, nerve trunks, cords, and peripheral nerve components ofthe brachial plexus.

In another embodiment, a sirtuin-modulating compound may be used totreat or prevent a polyglutamine disease. Exemplary polyglutaminediseases include Spinobulbar muscular atrophy (Kennedy disease),Huntington's Disease (HD), Dentatorubral-pallidoluysian atrophy (HawRiver syndrome), Spinocerebellar ataxia type 1, Spinocerebellar ataxiatype 2, Spinocerebellar ataxia type 3 (Machado-Joseph disease),Spinocerebellar ataxia type 6, Spinocerebellar ataxia type 7, andSpinocerebellar ataxia type 17.

In certain embodiments, the invention provides a method to treat acentral nervous system cell to prevent damage in response to a decreasein blood flow to the cell. Typically the severity of damage that may beprevented will depend in large part on the degree of reduction in bloodflow to the cell and the duration of the reduction. In certainembodiments, apoptotic or necrotic cell death may be prevented. In stilla further embodiment, ischemic-mediated damage, such as cytotoxic edemaor central nervous system tissue anoxemia, may be prevented. In eachembodiment, the central nervous system cell may be a spinal cell or abrain cell.

Another aspect encompasses administrating a sirtuin-modulating compoundto a subject to treat a central nervous system ischemic condition. Anumber of central nervous system ischemic conditions may be treated bythe sirtuin-modulating compounds described herein. In certainembodiments, the ischemic condition is a stroke that results in any typeof ischemic central nervous system damage, such as apoptotic or necroticcell death, cytotoxic edema or central nervous system tissue anoxia. Thestroke may impact any area of the brain or be caused by any etiologycommonly known to result in the occurrence of a stroke. In onealternative of this embodiment, the stroke is a brain stem stroke. Inanother alternative of this embodiment, the stroke is a cerebellarstroke. In still another embodiment, the stroke is an embolic stroke. Inyet another alternative, the stroke may be a hemorrhagic stroke. In afurther embodiment, the stroke is a thrombotic stroke.

In yet another aspect, a sirtuin-modulating compound may be administeredto reduce infarct size of the ischemic core following a central nervoussystem ischemic condition. Moreover, a sirtuin-modulating compound mayalso be beneficially administered to reduce the size of the ischemicpenumbra or transitional zone following a central nervous systemischemic condition.

In certain embodiments, a combination drug regimen may include drugs orcompounds for the treatment or prevention of neurodegenerative disordersor secondary conditions associated with these conditions. Thus, acombination drug regimen may include one or more sirtuin activators andone or more anti-neurodegeneration agents.

Blood Coagulation Disorders

In other aspects, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein can be used to treat or preventblood coagulation disorders (or hemostatic disorders). As usedinterchangeably herein, the terms “hemostasis”, “blood coagulation,” and“blood clotting” refer to the control of bleeding, including thephysiological properties of vasoconstriction and coagulation. Bloodcoagulation assists in maintaining the integrity of mammaliancirculation after injury, inflammation, disease, congenital defect,dysfunction or other disruption. Further, the formation of blood clotsdoes not only limit bleeding in case of an injury (hemostasis), but maylead to serious organ damage and death in the context of atheroscleroticdiseases by occlusion of an important artery or vein. Thrombosis is thusblood clot formation at the wrong time and place.

Accordingly, the present invention provides anticoagulation andantithrombotic treatments aiming at inhibiting the formation of bloodclots in order to prevent or treat blood coagulation disorders, such asmyocardial infarction, stroke, loss of a limb by peripheral arterydisease or pulmonary embolism.

As used interchangeably herein, “modulating or modulation of hemostasis”and “regulating or regulation of hemostasis” includes the induction(e.g., stimulation or increase) of hemostasis, as well as the inhibition(e.g., reduction or decrease) of hemostasis.

In one aspect, the invention provides a method for reducing orinhibiting hemostasis in a subject by administering a sirtuin-modulatingcompound that increases the level and/or activity of a sirtuin protein.The compositions and methods or uses disclosed herein are useful for thetreatment or prevention of thrombotic disorders. As used herein, theterm “thrombotic disorder” includes any disorder or conditioncharacterized by excessive or unwanted coagulation or hemostaticactivity, or a hypercoagulable state. Thrombotic disorders includediseases or disorders involving platelet adhesion and thrombusformation, and may manifest as an increased propensity to formthromboses, e.g., an increased number of thromboses, thrombosis at anearly age, a familial tendency towards thrombosis, and thrombosis atunusual sites.

In another embodiment, a combination drug regimen may include drugs orcompounds for the treatment or prevention of blood coagulation disordersor secondary conditions associated with these conditions. Thus, acombination drug regimen may include one or more sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteinand one or more anti-coagulation or anti-thrombosis agents.

Weight Control

In another aspect, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be used for treating orpreventing weight gain or obesity in a subject. For example,sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may be used, for example, to treat or preventhereditary obesity, dietary obesity, hormone related obesity, obesityrelated to the administration of medication, to reduce the weight of asubject, or to reduce or prevent weight gain in a subject. A subject inneed of such a treatment may be a subject who is obese, likely to becomeobese, overweight, or likely to become overweight. Subjects who arelikely to become obese or overweight can be identified, for example,based on family history, genetics, diet, activity level, medicationintake, or various combinations thereof.

In yet other embodiments, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be administered tosubjects suffering from a variety of other diseases and conditions thatmay be treated or prevented by promoting weight loss in the subject.Such diseases include, for example, high blood pressure, hypertension,high blood cholesterol, dyslipidemia, type 2 diabetes, insulinresistance, glucose intolerance, hyperinsulinemia, coronary heartdisease, angina pectoris, congestive heart failure, stroke, gallstones,cholecystitis and cholelithiasis, gout, osteoarthritis, obstructivesleep apnea and respiratory problems, some types of cancer (such asendometrial, breast, prostate, and colon), complications of pregnancy,poor female reproductive health (such as menstrual irregularities,infertility, irregular ovulation), bladder control problems (such asstress incontinence); uric acid nephrolithiasis; psychological disorders(such as depression, eating disorders, distorted body image, and lowself-esteem). Finally, patients with AIDS can develop lipodystrophy orinsulin resistance in response to combination therapies for AIDS.

In another embodiment, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used for inhibitingadipogenesis or fat cell differentiation, whether in vitro or in vivo.Such methods or uses may be used for treating or preventing obesity.

In other embodiments, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used for reducingappetite and/or increasing satiety, thereby causing weight loss oravoidance of weight gain. A subject in need of such a treatment may be asubject who is overweight, obese or a subject likely to becomeoverweight or obese. The method may comprise administering daily or,every other day, or once a week, a dose, e.g., in the form of a pill, toa subject. The dose may be an “appetite reducing dose.”

In an exemplary embodiment, sirtuin-modulating compounds that increasethe level and/or activity of a sirtuin protein may be administered as acombination therapy for treating or preventing weight gain or obesity.For example, one or more sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be administered incombination with one or more anti-obesity agents.

In another embodiment, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be administered to reducedrug-induced weight gain. For example, a sirtuin-modulating compoundthat increases the level and/or activity of a sirtuin protein may beadministered as a combination therapy with medications that maystimulate appetite or cause weight gain, in particular, weight gain dueto factors other than water retention.

Metabolic Disorders/Diabetes

In another aspect, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be used for treating orpreventing a metabolic disorder, such as insulin-resistance, apre-diabetic state, type II diabetes, and/or complications thereof.Administration of a sirtuin-modulating compound that increases the leveland/or activity of a sirtuin protein may increase insulin sensitivityand/or decrease insulin levels in a subject. A subject in need of such atreatment may be a subject who has insulin resistance or other precursorsymptom of type II diabetes, who has type II diabetes, or who is likelyto develop any of these conditions. For example, the subject may be asubject having insulin resistance, e.g., having high circulating levelsof insulin and/or associated conditions, such as hyperlipidemia,dyslipogenesis, hypercholesterolemia, impaired glucose tolerance, highblood glucose sugar level, other manifestations of syndrome X,hypertension, atherosclerosis and lipodystrophy.

In an exemplary embodiment, sirtuin-modulating compounds that increasethe level and/or activity of a sirtuin protein may be administered as acombination therapy for treating or preventing a metabolic disorder. Forexample, one or more sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be administered incombination with one or more anti-diabetic agents.

Inflammatory Diseases

In other aspects, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein can be used to treat or prevent adisease or disorder associated with inflammation. Sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteinmay be administered prior to the onset of, at, or after the initiationof inflammation. When used prophylactically, the compounds arepreferably provided in advance of any inflammatory response or symptom.Administration of the compounds may prevent or attenuate inflammatoryresponses or symptoms.

In another embodiment, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used to treat orprevent allergies and respiratory conditions, including asthma,bronchitis, pulmonary fibrosis, allergic rhinitis, oxygen toxicity,emphysema, chronic bronchitis, acute respiratory distress syndrome, andany chronic obstructive pulmonary disease (COPD). The compounds may beused to treat chronic hepatitis infection, including hepatitis B andhepatitis C.

Additionally, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be used to treat autoimmunediseases, and/or inflammation associated with autoimmune diseases, suchas arthritis, including rheumatoid arthritis, psoriatic arthritis, andankylosing spondylitis, as well as organ-tissue autoimmune diseases(e.g., Raynaud's syndrome), ulcerative colitis, Crohn's disease, oralmucositis, scleroderma, myasthenia gravis, transplant rejection,endotoxin shock, sepsis, psoriasis, eczema, dermatitis, multiplesclerosis, autoimmune thyroiditis, uveitis, systemic lupuserythematosis, Addison's disease, autoimmune polyglandular disease (alsoknown as autoimmune polyglandular syndrome), and Grave's disease.

In certain embodiments, one or more sirtuin-modulating compounds thatincrease the level and/or activity of a sirtuin protein may be takenalone or in combination with other compounds useful for treating orpreventing inflammation.

Flushing

In another aspect, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be used for reducing theincidence or severity of flushing and/or hot flashes which are symptomsof a disorder. For instance, the subject method includes the use ofsirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein, alone or in combination with other agents, forreducing incidence or severity of flushing and/or hot flashes in cancerpatients. In other embodiments, the method provides for the use ofsirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein to reduce the incidence or severity of flushing and/orhot flashes in menopausal and post-menopausal woman.

In another aspect, sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may be used as a therapy forreducing the incidence or severity of flushing and/or hot flashes whichare side-effects of another drug therapy, e.g., drug-induced flushing.In certain embodiments, a method for treating and/or preventingdrug-induced flushing comprises administering to a patient in needthereof a formulation comprising at least one flushing inducing compoundand at least one sirtuin-modulating compound that increases the leveland/or activity of a sirtuin protein. In other embodiments, a method fortreating drug induced flushing comprises separately administering one ormore compounds that induce flushing and one or more sirtuin-modulatingcompounds, e.g., wherein the sirtuin-modulating compound and flushinginducing agent have not been formulated in the same compositions. Whenusing separate formulations, the sirtuin-modulating compound may beadministered (1) at the same as administration of the flushing inducingagent, (2) intermittently with the flushing inducing agent, (3)staggered relative to administration of the flushing inducing agent, (4)prior to administration of the flushing inducing agent, (5) subsequentto administration of the flushing inducing agent, and (6) variouscombination thereof. Exemplary flushing inducing agents include, forexample, niacin, raloxifene, antidepressants, anti-psychotics,chemotherapeutics, calcium channel blockers, and antibiotics.

In certain embodiments, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used to reduceflushing side effects of a vasodilator or an antilipemic agent(including anticholesteremic agents and lipotropic agents). In anexemplary embodiment, a sirtuin-modulating compound that increases thelevel and/or activity of a sirtuin protein may be used to reduceflushing associated with the administration of niacin.

In another embodiment, the invention provides a method for treatingand/or preventing hyperlipidemia with reduced flushing side effects. Inanother representative embodiment, the method involves the use ofsirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein to reduce flushing side effects of raloxifene. Inanother representative embodiment, the method involves the use ofsirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein to reduce flushing side effects of antidepressants oranti-psychotic agent. For instance, sirtuin-modulating compounds thatincrease the level and/or activity of a sirtuin protein can be used inconjunction (administered separately or together) with a serotoninreuptake inhibitor, or a 5HT2 receptor antagonist.

In certain embodiments, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used as part of atreatment with a serotonin reuptake inhibitor (SRI) to reduce flushing.In still another representative embodiment, sirtuin-modulating compoundsthat increase the level and/or activity of a sirtuin protein may be usedto reduce flushing side effects of chemotherapeutic agents, such ascyclophosphamide and tamoxifen.

In another embodiment, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used to reduceflushing side effects of calcium channel blockers, such as amlodipine.

In another embodiment, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used to reduceflushing side effects of antibiotics. For example, sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteincan be used in combination with levofloxacin.

Ocular Disorders

One aspect of the present invention is a method for inhibiting, reducingor otherwise treating vision impairment by administering to a patient atherapeutic dosage of sirtuin modulator selected from a compounddisclosed herein, or a pharmaceutically acceptable salt, prodrug or ametabolic derivative thereof.

In certain aspects of the invention, the vision impairment is caused bydamage to the optic nerve or central nervous system. In particularembodiments, optic nerve damage is caused by high intraocular pressure,such as that created by glaucoma. In other particular embodiments, opticnerve damage is caused by swelling of the nerve, which is oftenassociated with an infection or an immune (e.g., autoimmune) responsesuch as in optic neuritis.

In certain aspects of the invention, the vision impairment is caused byretinal damage. In particular embodiments, retinal damage is caused bydisturbances in blood flow to the eye (e.g., arteriosclerosis,vasculitis). In particular embodiments, retinal damage is caused bydisruption of the macula (e.g., exudative or non-exudative maculardegeneration).

Exemplary retinal diseases include Exudative Age Related MacularDegeneration, Nonexudative Age Related Macular Degeneration, RetinalElectronic Prosthesis and RPE Transplantation Age Related MacularDegeneration, Acute Multifocal Placoid Pigment Epitheliopathy, AcuteRetinal Necrosis, Best Disease, Branch Retinal Artery Occlusion, BranchRetinal Vein Occlusion, Cancer Associated and Related AutoimmuneRetinopathies, Central Retinal Artery Occlusion, Central Retinal VeinOcclusion, Central Serous Chorioretinopathy, Eales Disease, EpimacularMembrane, Lattice Degeneration, Macroaneurysm, Diabetic Macular Edema,Irvine-Gass Macular Edema, Macular Hole, Subretinal NeovascularMembranes, Diffuse Unilateral Subacute Neuroretinitis, NonpseudophakicCystoid Macular Edema, Presumed Ocular Histoplasmosis Syndrome,Exudative Retinal Detachment, Postoperative Retinal Detachment,Proliferative Retinal Detachment, Rhegmatogenous Retinal Detachment,Tractional Retinal Detachment, Retinitis Pigmentosa, CMV Retinitis,Retinoblastoma, Retinopathy of Prematurity, Birdshot Retinopathy,Background Diabetic Retinopathy, Proliferative Diabetic Retinopathy,Hemoglobinopathies Retinopathy, Purtscher Retinopathy, ValsalvaRetinopathy, Juvenile Retinoschisis, Senile Retinoschisis, TersonSyndrome and White Dot Syndromes.

Other exemplary diseases include ocular bacterial infections (e.g.conjunctivitis, keratitis, tuberculosis, syphilis, gonorrhea), viralinfections (e.g., Ocular Herpes Simplex Virus, Varicella Zoster Virus,Cytomegalovirus retinitis, Human Immunodeficiency Virus (HIV)) as wellas progressive outer retinal necrosis secondary to HIV or otherHIV-associated and other immunodeficiency-associated ocular diseases. Inaddition, ocular diseases include fungal infections (e.g., Candidachoroiditis, histoplasmosis), protozoal infections (e.g., toxoplasmosis)and others such as ocular toxocariasis and sarcoidosis.

One aspect of the invention is a method for inhibiting, reducing ortreating vision impairment in a subject undergoing treatment with achemotherapeutic drug (e.g., a neurotoxic drug, or a drug that raisesintraocular pressure, such as a steroid), by administering to thesubject in need of such treatment a therapeutic dosage of a sirtuinmodulator disclosed herein.

Another aspect of the invention is a method for inhibiting, reducing ortreating vision impairment in a subject undergoing surgery, includingocular or other surgeries performed in the prone position such as spinalcord surgery, by administering to the subject in need of such treatmenta therapeutic dosage of a sirtuin modulator disclosed herein. Ocularsurgeries include cataract, iridotomy and lens replacements.

Another aspect of the invention is the treatment, including inhibitionand prophylactic treatment, of age related ocular diseases includecataracts, dry eye, age-related macular degeneration (AMD), retinaldamage and the like, by administering to the subject in need of suchtreatment a therapeutic dosage of a sirtuin modulator disclosed herein.

Another aspect of the invention is the prevention or treatment of damageto the eye caused by stress, chemical insult or radiation, byadministering to the subject in need of such treatment a therapeuticdosage of a sirtuin modulator disclosed herein. Radiation orelectromagnetic damage to the eye can include that caused by CRT's orexposure to sunlight or UV.

In certain embodiments, a combination drug regimen may include drugs orcompounds for the treatment or prevention of ocular disorders orsecondary conditions associated with these conditions. Thus, acombination drug regimen may include one or more sirtuin activators andone or more therapeutic agents for the treatment of an ocular disorder.

In certain embodiments, a sirtuin modulator can be administered inconjunction with a therapy for reducing intraocular pressure. In anotherembodiment, a sirtuin modulator can be administered in conjunction witha therapy for treating and/or preventing glaucoma. In yet anotherembodiment, a sirtuin modulator can be administered in conjunction witha therapy for treating and/or preventing optic neuritis. In certainembodiments, a sirtuin modulator can be administered in conjunction witha therapy for treating and/or preventing CMV Retinopathy. In anotherembodiment, a sirtuin modulator can be administered in conjunction witha therapy for treating and/or preventing multiple sclerosis.

Mitochondrial-Associated Diseases and Disorders

In certain embodiments, the invention provides methods or uses fortreating diseases or disorders that would benefit from increasedmitochondrial activity. The methods or uses involve administering to asubject in need thereof a therapeutically effective amount of asirtuin-modulating compound. Increased mitochondrial activity refers toincreasing activity of the mitochondria while maintaining the overallnumbers of mitochondria (e.g., mitochondrial mass), increasing thenumbers of mitochondria thereby increasing mitochondrial activity (e.g.,by stimulating mitochondrial biogenesis), or combinations thereof. Incertain embodiments, diseases and disorders that would benefit fromincreased mitochondrial activity include diseases or disordersassociated with mitochondrial dysfunction.

In certain embodiments, methods or uses for treating diseases ordisorders that would benefit from increased mitochondrial activity maycomprise identifying a subject suffering from a mitochondrialdysfunction. Methods or uses for diagnosing a mitochondrial dysfunctionmay involve molecular genetics, pathologic and/or biochemical analyses.Diseases and disorders associated with mitochondrial dysfunction includediseases and disorders in which deficits in mitochondrial respiratorychain activity contribute to the development of pathophysiology of suchdiseases or disorders in a mammal. Diseases or disorders that wouldbenefit from increased mitochondrial activity generally include forexample, diseases in which free radical mediated oxidative injury leadsto tissue degeneration, diseases in which cells inappropriately undergoapoptosis, and diseases in which cells fail to undergo apoptosis.

In certain embodiments, the invention provides methods or uses fortreating a disease or disorder that would benefit from increasedmitochondrial activity that involves administering to a subject in needthereof one or more sirtuin-modulating compounds in combination withanother therapeutic agent such as, for example, an agent useful fortreating mitochondrial dysfunction or an agent useful for reducing asymptom associated with a disease or disorder involving mitochondrialdysfunction.

In exemplary embodiments, the invention provides methods or uses fortreating diseases or disorders that would benefit from increasedmitochondrial activity by administering to a subject a therapeuticallyeffective amount of a sirtuin-modulating compound. Exemplary diseases ordisorders include, for example, neuromuscular disorders (e.g.,Friedreich's Ataxia, muscular dystrophy, multiple sclerosis, etc.),disorders of neuronal instability (e.g., seizure disorders, migraine,etc.), developmental delay, neurodegenerative disorders (e.g.,Alzheimer's Disease, Parkinson's Disease, amyotrophic lateral sclerosis,etc.), ischemia, renal tubular acidosis, age-related neurodegenerationand cognitive decline, chemotherapy fatigue, age-related orchemotherapy-induced menopause or irregularities of menstrual cycling orovulation, mitochondrial myopathies, mitochondrial damage (e.g., calciumaccumulation, excitotoxicity, nitric oxide exposure, hypoxia, etc.), andmitochondrial deregulation.

Muscular dystrophy refers to a family of diseases involvingdeterioration of neuromuscular structure and function, often resultingin atrophy of skeletal muscle and myocardial dysfunction, such asDuchenne muscular dystrophy. In certain embodiments, sirtuin-modulatingcompounds may be used for reducing the rate of decline in muscularfunctional capacities and for improving muscular functional status inpatients with muscular dystrophy.

In certain embodiments, sirtuin-modulating compounds may be useful fortreatment mitochondrial myopathies. Mitochondrial myopathies range frommild, slowly progressive weakness of the extraocular muscles to severe,fatal infantile myopathies and multisystem encephalomyopathies. Somesyndromes have been defined, with some overlap between them. Establishedsyndromes affecting muscle include progressive external ophthalmoplegia,the Kearns-Sayre syndrome (with ophthalmoplegia, pigmentary retinopathy,cardiac conduction defects, cerebellar ataxia, and sensorineuraldeafness), the MELAS syndrome (mitochondrial encephalomyopathy, lacticacidosis, and stroke-like episodes), the MERFF syndrome (myoclonicepilepsy and ragged red fibers), limb-girdle distribution weakness, andinfantile myopathy (benign or severe and fatal).

In certain embodiments, sirtuin-modulating compounds may be useful fortreating patients suffering from toxic damage to mitochondria, such as,toxic damage due to calcium accumulation, excitotoxicity, nitric oxideexposure, drug induced toxic damage, or hypoxia.

In certain embodiments, sirtuin-modulating compounds may be useful fortreating diseases or disorders associated with mitochondrialderegulation.

Muscle Performance

In other embodiments, the invention provides methods or uses forenhancing muscle performance by administering a therapeuticallyeffective amount of a sirtuin-modulating compound. For example,sirtuin-modulating compounds may be useful for improving physicalendurance (e.g., ability to perform a physical task such as exercise,physical labor, sports activities, etc.), inhibiting or retardingphysical fatigues, enhancing blood oxygen levels, enhancing energy inhealthy individuals, enhance working capacity and endurance, reducingmuscle fatigue, reducing stress, enhancing cardiac and cardiovascularfunction, improving sexual ability, increasing muscle ATP levels, and/orreducing lactic acid in blood. In certain embodiments, the methods oruses involve administering an amount of a sirtuin-modulating compoundthat increase mitochondrial activity, increase mitochondrial biogenesis,and/or increase mitochondrial mass.

Sports performance refers to the ability of the athlete's muscles toperform when participating in sports activities. Enhanced sportsperformance, strength, speed and endurance are measured by an increasein muscular contraction strength, increase in amplitude of musclecontraction, shortening of muscle reaction time between stimulation andcontraction. Athlete refers to an individual who participates in sportsat any level and who seeks to achieve an improved level of strength,speed and endurance in their performance, such as, for example, bodybuilders, bicyclists, long distance runners, short distance runners,etc. Enhanced sports performance in manifested by the ability toovercome muscle fatigue, ability to maintain activity for longer periodsof time, and have a more effective workout.

In the arena of athlete muscle performance, it is desirable to createconditions that permit competition or training at higher levels ofresistance for a prolonged period of time.

It is contemplated that the methods or uses of the present inventionwill also be effective in the treatment of muscle related pathologicalconditions, including acute sarcopenia, for example, muscle atrophyand/or cachexia associated with burns, bed rest, limb immobilization, ormajor thoracic, abdominal, and/or orthopedic surgery.

In certain embodiments, the invention provides novel dietarycompositions comprising sirtuin modulators, a method for theirpreparation, and a method of using the compositions for improvement ofsports performance. Accordingly, provided are therapeutic compositions,foods and beverages that have actions of improving physical enduranceand/or inhibiting physical fatigues for those people involved inbroadly-defined exercises including sports requiring endurance andlabors requiring repeated muscle exertions. Such dietary compositionsmay additional comprise electrolytes, caffeine, vitamins, carbohydrates,etc.

Other Uses

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may be used for treating or preventing viralinfections (such as infections by influenza, herpes or papilloma virus)or as antifungal agents. In certain embodiments, sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteinmay be administered as part of a combination drug therapy with anothertherapeutic agent for the treatment of viral diseases. In anotherembodiment, sirtuin-modulating compounds that increase the level and/oractivity of a sirtuin protein may be administered as part of acombination drug therapy with another anti-fungal agent.

Subjects that may be treated as described herein include eukaryotes,such as mammals, e.g., humans, ovines, bovines, equines, porcines,canines, felines, non-human primate, mice, and rats. Cells that may betreated include eukaryotic cells, e.g., from a subject described above,or plant cells, yeast cells and prokaryotic cells, e.g., bacterialcells. For example, modulating compounds may be administered to farmanimals to improve their ability to withstand farming conditions longer.

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may also be used to increase lifespan, stressresistance, and resistance to apoptosis in plants. In certainembodiments, a compound is applied to plants, e.g., on a periodic basis,or to fungi. In another embodiment, plants are genetically modified toproduce a compound. In another embodiment, plants and fruits are treatedwith a compound prior to picking and shipping to increase resistance todamage during shipping. Plant seeds may also be contacted with compoundsdescribed herein, e.g., to preserve them.

In other embodiments, sirtuin-modulating compounds that increase thelevel and/or activity of a sirtuin protein may be used for modulatinglifespan in yeast cells. Situations in which it may be desirable toextend the lifespan of yeast cells include any process in which yeast isused, e.g., the making of beer, yogurt, and bakery items, e.g., bread.Use of yeast having an extended lifespan can result in using less yeastor in having the yeast be active for longer periods of time. Yeast orother mammalian cells used for recombinantly producing proteins may alsobe treated as described herein.

Sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein may also be used to increase lifespan, stressresistance and resistance to apoptosis in insects. In this embodiment,compounds would be applied to useful insects, e.g., bees and otherinsects that are involved in pollination of plants. In a specificembodiment, a compound would be applied to bees involved in theproduction of honey. Generally, the methods or uses described herein maybe applied to any organism, e.g., eukaryote, which may have commercialimportance. For example, they can be applied to fish (aquaculture) andbirds (e.g., chicken and fowl).

Higher doses of sirtuin-modulating compounds that increase the leveland/or activity of a sirtuin protein may also be used as a pesticide byinterfering with the regulation of silenced genes and the regulation ofapoptosis during development. In this embodiment, a compound may beapplied to plants using a method known in the art that ensures thecompound is bio-available to insect larvae, and not to plants.

At least in view of the link between reproduction and longevity,sirtuin-modulating compounds that increase the level and/or activity ofa sirtuin protein can be applied to affect the reproduction of organismssuch as insects, animals and microorganisms.

Additional Embodiments

In one aspect, the present invention relates to a method of increasingsirtuin-1 activity in a cell comprising the step of contacting the cellwith a compound of Formula (I) or a pharmaceutically acceptable salt orcorresponding pharmaceutical composition, respectively, thereof.

In one aspect, the present invention relates to a method for treatinginsulin resistance, a metabolic syndrome, diabetes, or complicationsthereof, or for increasing insulin sensitivity, comprising administeringa compound or a pharmaceutically acceptable salt or correspondingpharmaceutical composition, respectively, thereof, to a subject in needthereof.

In one aspect, the present invention relates to a method for treatingmetabolic dysfunctions comprising administering a compound or apharmaceutically acceptable salt or corresponding pharmaceuticalcomposition, respectively, thereof, to a subject in need thereof.

In one aspect, the present invention relates to a method for treatingdiseases or disorders resulting from diminished SIRT1 expression oractivity, which comprises administering a compound or a pharmaceuticallyacceptable salt or corresponding pharmaceutical composition,respectively, thereof, to a subject in need thereof.

In one aspect, the present invention relates to a method where thediseases or disorders resulting from diminished SIRT1 expression oractivity are selected from, but not limited to aging or stress,diabetes, metabolic dysfunctions, neurodegenerative diseases,cardiovascular disease, cancer or inflammatory disease.

In one aspect, the present invention relates to a method, where diseasesrelated to aging or stress, diabetes, metabolic dysfunctions,neurodegenerative diseases, cardiovascular disease, cancer orinflammatory disease are selected from psoriasis, atopic dermatitis,acne, rosacea, inflammatory bowel disease, osteoporosis, sepsis,arthritis, COPD, systemic lupus erythematosus and ophthalmicinflammation.

In one aspect, the present invention relates to a method, where diseasesrelated to aging or stress, diabetes, metabolic dysfunctions,neurodegenerative diseases, cardiovascular disease, cancer orinflammatory disease are selected from psoriasis, atopic dermatitis,acne, rosacea, inflammatory bowel disease, osteoporosis, sepsis,arthritis, COPD, systemic lupus erythematosus and ophthalmicinflammation.

In one aspect, the present invention relates to a method for treatingpsoriasis, which comprises administering a compound or apharmaceutically acceptable salt or corresponding pharmaceuticalcomposition, respectively, thereof, to a subject in need thereof.

In one aspect, the present invention relates to administering a compoundor a pharmaceutically acceptable salt or corresponding pharmaceuticalcomposition, respectively, thereof, for use in therapy in treating asubject suffering from or susceptible to insulin resistance, a metabolicsyndrome, diabetes, or complications thereof, or for increasing insulinsensitivity in a subject.

In one aspect, the present invention relates to a use of administering acompound or a pharmaceutically acceptable salt or correspondingpharmaceutical composition, respectively, thereof, in the manufacture ofa medicament for use in the treatment of insulin resistance, a metabolicsyndrome, diabetes, or complications thereof, or for increasing insulinsensitivity in a subject.

Assays

Yet other methods or uses contemplated herein include screening methodsfor identifying compounds or agents that modulate sirtuins. An agent maybe a nucleic acid, such as an aptamer. Assays may be conducted in a cellbased or cell free format. For example, an assay may comprise incubating(or contacting) a sirtuin with a test agent under conditions in which asirtuin can be modulated by an agent known to modulate the sirtuin, andmonitoring or determining the level of modulation of the sirtuin in thepresence of the test agent relative to the absence of the test agent.The level of modulation of a sirtuin can be determined by determiningits ability to deacetylate a substrate. Exemplary substrates areacetylated peptides which can be obtained from BIOMOL (Plymouth Meeting,Pa.). Preferred substrates include peptides of p53, such as thosecomprising an acetylated K382. A particularly preferred substrate is theFluor de Lys-SIRT1 (BIOMOL), i.e., the acetylated peptideArg-His-Lys-Lys. Other substrates are peptides from human histones H3and H4 or an acetylated amino acid. Substrates may be fluorogenic. Thesirtuin may be SIRT1, Sir2, SIRT3, or a portion thereof. For example,recombinant SIRT1 can be obtained from BIOMOL. The reaction may beconducted for about 30 minutes and stopped, e.g., with nicotinamide. TheHDAC fluorescent activity assay/drug discovery kit (AK-500, BIOMOLResearch Laboratories) may be used to determine the level ofacetylation. Similar assays are described in Bitterman et al. (2002) J.Biol. Chem. 277:45099. The level of modulation of the sirtuin in anassay may be compared to the level of modulation of the sirtuin in thepresence of one or more (separately or simultaneously) compoundsdescribed herein, which may serve as positive or negative controls.Sirtuins for use in the assays may be full length sirtuin proteins orportions thereof. Since it has been shown herein that activatingcompounds appear to interact with the N-terminus of SIRT1, proteins foruse in the assays include N-terminal portions of sirtuins, e.g., aboutamino acids 1-176 or 1-255 of SIRT1; about amino acids 1-174 or 1-252 ofSir2.

In certain embodiments, a screening assay comprises (i) contacting asirtuin with a test agent and an acetylated substrate under conditionsappropriate for the sirtuin to deacetylate the substrate in the absenceof the test agent; and (ii) determining the level of acetylation of thesubstrate, wherein a lower level of acetylation of the substrate in thepresence of the test agent relative to the absence of the test agentindicates that the test agent stimulates deacetylation by the sirtuin,whereas a higher level of acetylation of the substrate in the presenceof the test agent relative to the absence of the test agent indicatesthat the test agent inhibits deacetylation by the sirtuin.

In another embodiment, the screening assay may detect the formation of a2′/3′-O-acetyl-ADP-ribose product of sirtuin-mediated NAD-dependentdeacetylation. This O-acetyl-ADP-ribose product is formed in equimolarquantities with the deacetylated peptide product of the sirtuindeacetylation reaction. Accordingly, the screening assay may include (i)contacting a sirtuin with a test agent and an acetylated substrate underconditions appropriate for the sirtuin to deacetylate the substrate inthe absence of the test agent; and (ii) determining the amount ofO-acetyl-ADP-ribose formation, wherein an increase inO-acetyl-ADP-ribose formation in the presence of the test agent relativeto the absence of the test agent indicates that the test agentstimulates deacetylation by the sirtuin, while a decrease inO-acetyl-ADP-ribose formation in the presence of the test agent relativeto the absence of the test agent indicates that the test agent inhibitsdeacetylation by the sirtuin.

Methods or uses for identifying an agent that modulates, e.g.,stimulates, sirtuins in vivo may comprise (i) contacting a cell with atest agent and a substrate that is capable of entering a cell in thepresence of an inhibitor of class I and class II HDACs under conditionsappropriate for the sirtuin to deacetylate the substrate in the absenceof the test agent; and (ii) determining the level of acetylation of thesubstrate, wherein a lower level of acetylation of the substrate in thepresence of the test agent relative to the absence of the test agentindicates that the test agent stimulates deacetylation by the sirtuin,whereas a higher level of acetylation of the substrate in the presenceof the test agent relative to the absence of the test agent indicatesthat the test agent inhibits deacetylation by the sirtuin. A preferredsubstrate is an acetylated peptide, which is also preferablyfluorogenic, as further described herein. The method may furthercomprise lysing the cells to determine the level of acetylation of thesubstrate. Substrates may be added to cells at a concentration rangingfrom about 1 μM to about 10 mM, preferably from about 10 μM to 1 mM,even more preferably from about 100 μM to 1 mM, such as about 200 μM. Apreferred substrate is an acetylated lysine, e.g., 6-acetyl lysine(Fluor de Lys, FdL) or Fluor de Lys-SIRT1. A preferred inhibitor ofclass I and class II HDACs is trichostatin A (TSA), which may be used atconcentrations ranging from about 0.01 to 100 μM, preferably from about0.1 to 10 μM, such as 1 μM. Incubation of cells with the test compoundand the substrate may be conducted for about 10 minutes to 5 hours,preferably for about 1-3 hours. Since TSA inhibits all class I and classII HDACs, and that certain substrates, e.g., Fluor de Lys, is a poorsubstrate for SIRT2 and even less a substrate for SIRT3-7, such an assaymay be used to identify modulators of SIRT1 in vivo.

Methods

The present invention also relates to methods or uses for using SirtuinModulator compounds as defined herein in treating and/or preventing awide variety of diseases and disorders, which include, but are notlimited to, for example, diseases or disorders related to aging orstress, diabetes, obesity, neurodegenerative diseases, cardiovasculardisease, blood clotting disorders, inflammation, cancer, and/or flushingas well as diseases or disorders that would benefit from increasedmitochondrial activity, further which may be selected from or include,but are not limited to psoriasis, atopic dermatitis, acne, rosacea,inflammatory bowel disease, osteoporosis, sepsis, arthritis, COPD,systemic lupus erythematosus and ophthalmic inflammation.

In another aspect, the invention provides methods or uses for usingsirtuin-modulating compounds, or compositions comprisingsirtuin-modulating compounds. In certain embodiments, sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteinmay be used for a variety of therapeutic applications including, forexample, increasing the lifespan of a cell, and treating and/orpreventing a wide variety of diseases and disorders including, forexample, diseases or disorders related to aging or stress, diabetes,obesity, neurodegenerative diseases, chemotherapeutic-inducedneuropathy, neuropathy associated with an ischemic event, oculardiseases and/or disorders, cardiovascular disease, blood clottingdisorders, inflammation, and/or flushing, etc. Sirtuin-modulatingcompounds that increase the level and/or activity of a sirtuin proteinmay also be used for treating a disease or disorder in a subject thatwould benefit from increased mitochondrial activity, for enhancingmuscle performance, for increasing muscle ATP levels, or for treating orpreventing muscle tissue damage associated with hypoxia or ischemia. Inother embodiments, sirtuin-modulating compounds that decrease the leveland/or activity of a sirtuin protein may be used for a variety oftherapeutic applications including, for example, increasing cellularsensitivity to stress, increasing apoptosis, treatment of cancer,stimulation of appetite, and/or stimulation of weight gain, etc. Asdescribed further below, the methods or uses comprise administering to asubject in need thereof a pharmaceutically effective amount of asirtuin-modulating compound.

In certain aspects, the sirtuin-modulating compounds may be administeredalone or in combination with other compounds, including othersirtuin-modulating compounds, or other therapeutic agents.

In another aspect, the present invention relates to a method ofincreasing sirtuin-1 activity in a cell, which comprises the step ofcontacting the cell with a compound of Formulas (I) to (IV),corresponding anlogs or derivatives thereof (i.e., with hydrogensubstitution at the R² position) or a pharmaceutical acceptable saltthereof of the present invention.

In another aspect, the present invention relates to a method ofincreasing sirtuin-1 activity in a cell comprising the step ofcontacting the cell with a pharmaceutical composition of the presentinvention as defined herein

In another aspect, the present invention relates to a method fortreating insulin resistance, a metabolic syndrome, diabetes, orcomplications thereof, or for increasing insulin sensitivity, whichcomprises administering a compound a compound of Formulas (I) to (IV),corresponding anlogs or derivatives thereof (i.e., with hydrogensubstitution at the R² position) of the present invention to a subjectin need thereof.

In another aspect, the present invention relates to a method fortreating a subject suffering from or susceptible to insulin resistance,a metabolic syndrome, diabetes, or complications thereof, or forincreasing insulin sensitivity in a subject, comprising administering apharmaceutical composition of the present invention to the subject inneed thereof.

In another aspect, the present invention relates to a method fortreating insulin resistance, a metabolic syndrome, diabetes, orcomplications thereof, or for increasing insulin sensitivity, comprisingadministering a pharmaceutical composition of the present invention to asubject in need thereof.

In another aspect, the present invention relates to a method ofincreasing sirtuin-1 activity in a cell, which comprises the step ofcontacting a cell with a compound of Formulas (I) to (IV), correspondinganlogs or derivatives thereof (i.e., with hydrogen substitution at theR² position) or a pharmaceutical acceptable salt thereof.

In another aspect, the present invention relates to a method ofincreasing sirtuin-1 activity in a cell, which comprises the step ofcontacting a cell with a pharmaceutical composition of the presentinvention

In another aspect, the present invention relates to a method fortreating metabolic dysfunctions, which comprises administering acompound of Formulas (I) to (IV), corresponding anlogs or derivativesthereof (i.e., with hydrogen substitution at the R² position) or apharmaceutical acceptable salt thereof to a subject in need thereof.

In another aspect, the present invention relates to a method fortreating metabolic dysfunctions comprising administering apharmaceutical composition of the present invention to a subject in needthereof.

In another aspect, the present invention relates to a method fortreating diseases or disorders resulting from diminished SIRT1expression or activity, which comprises administering a compound ofFormulas (I) to (IV), corresponding anlogs or derivatives thereof (i.e.,with hydrogen substitution at the R² position) or a pharmaceuticalacceptable salt thereof to a subject in need thereof.

In another aspect, the present invention relates to method where thediseases or disorders resulting from diminished SIRT1 expression oractivity are selected from, but not limited to aging or stress,diabetes, metabolic dysfunctions, neurodegenerative diseases,cardiovascular disease, cancer or inflammatory disease.

In another aspect, the present invention relates to a method wherediseases related to aging or stress, diabetes, metabolic dysfunctions,neurodegenerative diseases, cardiovascular disease, cancer orinflammatory disease are selected from psoriasis, atopic dermatitis,acne, rosacea, inflammatory bowel disease, osteoporosis, sepsis,arthritis, COPD, systemic lupus erythematosus and ophthalmicinflammation.

In another aspect, the present invention relates to a method fortreating psoriasis, which comprises administering a compound of Formulas(I) to (IV), corresponding anlogs or derivatives thereof (i.e., withhydrogen substitution at the R² position) or a pharmaceutical acceptablesalt thereof to a subject in need thereof.

In another aspect, the present invention relates to a method fortreating psoriasis, which comprises administering a pharmaceuticalcomposition of the present invention to a subject in need thereof

Pharmaceutical Compositions and Formulations

In general, the present invention relates to substituted bridged ureaanalog compounds of Formulas (I) to (IV), corresponding anlogs orderivatives thereof (i.e., with hydrogen substitution at the R²position), or pharmaceutically acceptable salts thereof, correspondingpharmaceutical compositions, processes for making and use of suchcompounds, alone or in combination with other therapeutic agents, asSirtuin Modulators useful for increasing lifespan of a cell, and intreating and/or preventing a wide variety of diseases and disorders,which include, but are not limited to, for example, diseases ordisorders related to aging or stress, diabetes, obesity,neurodegenerative diseases, cardiovascular disease, blood clottingdisorders, inflammation, cancer, and/or flushing as well as diseases ordisorders that would benefit from increased mitochondrial activity.

In particular, the present invention relates to novel compounds ofFormulas (I) to (IV), corresponding anlogs or derivatives thereof (i.e.,with hydrogen substitution at the R² position) or a pharmaceuticalacceptable salt thereof and corresponding pharmaceutical compositionscomprising compounds of Formulas (I) to (IV), respectively.

In another aspect, the present invention relates to a pharmaceuticalcomposition comprising a pharmaceutically acceptable carrier or diluentand a compound of Formulas (I) to (IV), corresponding anlogs orderivatives thereof (i.e., with hydrogen substitution at the R²position) or a pharmaceutical acceptable salt thereof.

In another aspect, the present invention relates to a pharmaceuticalcomposition of the present invention, further comprising an additionalactive agent.

In another aspect, the present invention relates to a pharmaceuticalcomposition comprising a compound of Formulas (I) to (IV), correspondinganlogs or derivatives thereof (i.e., with hydrogen substitution at theR² position) or a pharmaceutical acceptable salt thereof and at leastone pharmaceutically acceptable carrier.

The compounds described herein may be formulated in a conventionalmanner using one or more physiologically or pharmaceutically acceptablecarriers or excipients. For example, compounds and theirpharmaceutically acceptable salts and solvates may be formulated foradministration by, for example, injection (e.g. SubQ, IM, IP),inhalation or insufflation (either through the mouth or the nose) ororal, buccal, sublingual, transdermal, nasal, parenteral or rectaladministration. In certain embodiments, a compound may be administeredlocally, at the site where the target cells are present, i.e., in aspecific tissue, organ, or fluid (e.g., blood, cerebrospinal fluid,etc.).

The compounds can be formulated for a variety of modes ofadministration, including systemic and topical or localizedadministration. Techniques and formulations generally may be found inRemington's Pharmaceutical Sciences, Meade Publishing Co., Easton, Pa.For parenteral administration, injection is preferred, includingintramuscular, intravenous, intraperitoneal, and subcutaneous. Forinjection, the compounds can be formulated in liquid solutions,preferably in physiologically compatible buffers such as Hank's solutionor Ringer's solution. In addition, the compounds may be formulated insolid form and redissolved or suspended immediately prior to use.Lyophilized forms are also included.

For oral administration, the pharmaceutical compositions may take theform of, for example, tablets, lozenges, or capsules prepared byconventional means with pharmaceutically acceptable excipients such asbinding agents (e.g., pregelatinized maize starch, polyvinylpyrrolidoneor hydroxypropyl methylcellulose); fillers (e.g., lactose,microcrystalline cellulose or calcium hydrogen phosphate); lubricants(e.g., magnesium stearate, talc or silica); disintegrants (e.g., potatostarch or sodium starch glycolate); or wetting agents (e.g., sodiumlauryl sulphate). The tablets may be coated by methods well known in theart. Liquid preparations for oral administration may take the form of,for example, solutions, syrups or suspensions, or they may be presentedas a dry product for constitution with water or other suitable vehiclebefore use. Such liquid preparations may be prepared by conventionalmeans with pharmaceutically acceptable additives such as suspendingagents (e.g., sorbitol syrup, cellulose derivatives or hydrogenatededible fats); emulsifying agents (e.g., lecithin or acacia); non-aqueousvehicles (e.g., almond oil, oily esters, ethyl alcohol or fractionatedvegetable oils); and preservatives (e.g., methyl orpropyl-p-hydroxybenzoates or sorbic acid). The preparations may alsocontain buffer salts, flavoring, coloring and sweetening agents asappropriate. Preparations for oral administration may be suitablyformulated to give controlled release of the active compound.

For administration by inhalation (e.g., pulmonary delivery), thecompounds may be conveniently delivered in the form of an aerosol spraypresentation from pressurized packs or a nebulizer, with the use of asuitable propellant, e.g., dichlorodifluoromethane,trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide orother suitable gas. In the case of a pressurized aerosol the dosage unitmay be determined by providing a valve to deliver a metered amount.Capsules and cartridges of, e.g., gelatin, for use in an inhaler orinsufflator may be formulated containing a powder mix of the compoundand a suitable powder base such as lactose or starch.

The compounds may be formulated for parenteral administration byinjection, e.g., by bolus injection or continuous infusion. Formulationsfor injection may be presented in unit dosage form, e.g., in ampoules orin multi-dose containers, with an added preservative. The compositionsmay take such forms as suspensions, solutions or emulsions in oily oraqueous vehicles, and may contain formulatory agents such as suspending,stabilizing and/or dispersing agents. Alternatively, the activeingredient may be in powder form for constitution with a suitablevehicle, e.g., sterile pyrogen-free water, before use.

The compounds may also be formulated in rectal compositions such assuppositories or retention enemas, e.g., containing conventionalsuppository bases such as cocoa butter or other glycerides.

In addition to the formulations described previously, compounds may alsobe formulated as a depot preparation. Such long acting formulations maybe administered by implantation (for example subcutaneously orintramuscularly) or by intramuscular injection. Thus, for example,compounds may be formulated with suitable polymeric or hydrophobicmaterials (for example as an emulsion in an acceptable oil) or ionexchange resins, or as sparingly soluble derivatives, for example, as asparingly soluble salt. Controlled release formula also includespatches.

In certain embodiments, the compounds described herein can be formulatedfor delivery to the central nervous system (CNS) (reviewed in Begley,Pharmacology & Therapeutics 104: 29-45 (2004)). Conventional approachesfor drug delivery to the CNS include: neurosurgical strategies (e.g.,intracerebral injection or intracerebroventricular infusion); molecularmanipulation of the agent (e.g., production of a chimeric fusion proteinthat comprises a transport peptide that has an affinity for anendothelial cell surface molecule in combination with an agent that isitself incapable of crossing the BBB) in an attempt to exploit one ofthe endogenous transport pathways of the BBB; pharmacological strategiesdesigned to increase the lipid solubility of an agent (e.g., conjugationof water-soluble agents to lipid or cholesterol carriers); and thetransitory disruption of the integrity of the BBB by hyperosmoticdisruption (resulting from the infusion of a mannitol solution into thecarotid artery or the use of a biologically active agent such as anangiotensin peptide).

Liposomes are a further drug delivery system which is easily injectable.Accordingly, in the method of invention the active compounds can also beadministered in the form of a liposome delivery system. Liposomes arewell known by those skilled in the art. Liposomes can be formed from avariety of phospholipids, such as cholesterol, stearylamine ofphosphatidylcholines. Liposomes usable for the method of inventionencompass all types of liposomes including, but not limited to, smallunilamellar vesicles, large unilamellar vesicles and multilamellarvesicles.

Another way to produce a formulation, particularly a solution, of acompound described herein, is through the use of cyclodextrin. Bycyclodextrin is meant α-, β-, or γ-cyclodextrin. Cyclodextrins aredescribed in detail in Pitha et al., U.S. Pat. No. 4,727,064.Cyclodextrins are cyclic oligomers of glucose; these compounds forminclusion complexes with any drug whose molecule can fit into thelipophile-seeking cavities of the cyclodextrin molecule.

Rapidly disintegrating or dissolving dosage forms are useful for therapid absorption, particularly buccal and sublingual absorption, ofpharmaceutically active agents. Fast melt dosage forms are beneficial topatients, such as aged and pediatric patients, who have difficulty inswallowing typical solid dosage forms, such as caplets and tablets.Additionally, fast melt dosage forms circumvent drawbacks associatedwith, for example, chewable dosage forms, wherein the length of time anactive agent remains in a patient's mouth plays an important role indetermining the amount of taste masking and the extent to which apatient may object to throat grittiness of the active agent.

Pharmaceutical compositions (including cosmetic preparations) maycomprise from about 0.00001 to 100% such as from 0.001 to 10% or from0.1% to 5% by weight of one or more compounds described herein. In otherembodiments, the pharmaceutical composition comprises: (i) 0.05 to 1000mg of the compounds of the invention, or a pharmaceutically acceptablesalt thereof, and (ii) 0.1 to 2 grams of one or more pharmaceuticallyacceptable excipients.

In some embodiments, a compound described herein is incorporated into atopical formulation containing a topical carrier that is generallysuited to topical drug administration and comprising any such materialknown in the art. The topical carrier may be selected so as to providethe composition in the desired form, e.g., as an ointment, lotion,cream, microemulsion, gel, oil, solution, or the like, and may becomprised of a material of either naturally occurring or syntheticorigin. It is preferable that the selected carrier not adversely affectthe active agent or other components of the topical formulation.Examples of suitable topical carriers for use herein include water,alcohols and other nontoxic organic solvents, glycerin, mineral oil,silicone, petroleum jelly, lanolin, fatty acids, vegetable oils,parabens, waxes, and the like.

Formulations may be colorless, odorless ointments, lotions, creams,microemulsions and gels.

The compounds may be incorporated into ointments, which generally aresemisolid preparations which are typically based on petrolatum or otherpetroleum derivatives. The specific ointment base to be used, as will beappreciated by those skilled in the art, is one that will provide foroptimum drug delivery, and, preferably, will provide for other desiredcharacteristics as well, e.g., emolliency or the like. As with othercarriers or vehicles, an ointment base should be inert, stable,nonirritating and nonsensitizing.

The compounds may be incorporated into lotions, which generally arepreparations to be applied to the skin surface without friction, and aretypically liquid or semiliquid preparations in which solid particles,including the active agent, are present in a water or alcohol base.Lotions are usually suspensions of solids, and may comprise a liquidoily emulsion of the oil-in-water type.

The compounds may be incorporated into creams, which generally areviscous liquid or semisolid emulsions, either oil-in-water orwater-in-oil. Cream bases are water-washable, and contain an oil phase,an emulsifier and an aqueous phase. The oil phase is generally comprisedof petrolatum and a fatty alcohol such as cetyl or stearyl alcohol; theaqueous phase usually, although not necessarily, exceeds the oil phasein volume, and generally contains a humectant. The emulsifier in a creamformulation, as explained in Remington's, supra, is generally anonionic, anionic, cationic or amphoteric surfactant.

The compounds may be incorporated into microemulsions, which generallyare thermodynamically stable, isotropically clear dispersions of twoimmiscible liquids, such as oil and water, stabilized by an interfacialfilm of surfactant molecules (Encyclopedia of Pharmaceutical Technology(New York: Marcel Dekker, 1992), volume 9).

The compounds may be incorporated into gel formulations, which generallyare semisolid systems consisting of either suspensions made up of smallinorganic particles (two-phase systems) or large organic moleculesdistributed substantially uniformly throughout a carrier liquid (singlephase gels). Although gels commonly employ aqueous carrier liquid,alcohols and oils can be used as the carrier liquid as well.

Other active agents may also be included in formulations, e.g., otheranti-inflammatory agents, analgesics, antimicrobial agents, antifungalagents, antibiotics, vitamins, antioxidants, and sunblock agentscommonly found in sunscreen formulations including, but not limited to,anthranilates, benzophenones (particularly benzophenone-3), camphorderivatives, cinnamates (e.g., octyl methoxycinnamate), dibenzoylmethanes (e.g., butyl methoxydibenzoyl methane), p-aminobenzoic acid(PABA) and derivatives thereof, and salicylates (e.g., octylsalicylate).

In certain topical formulations, the active agent is present in anamount in the range of approximately 0.25 wt. % to 75 wt. % of theformulation, preferably in the range of approximately 0.25 wt. % to 30wt. % of the formulation, more preferably in the range of approximately0.5 wt. % to 15 wt. % of the formulation, and most preferably in therange of approximately 1.0 wt. % to 10 wt. % of the formulation.

Conditions of the eye can be treated or prevented by, e.g., systemic,topical, intraocular injection of a compound, or by insertion of asustained release device that releases a compound. A compound may bedelivered in a pharmaceutically acceptable ophthalmic vehicle, such thatthe compound is maintained in contact with the ocular surface for asufficient time period to allow the compound to penetrate the cornealand internal regions of the eye, as for example the anterior chamber,posterior chamber, vitreous body, aqueous humor, vitreous humor, cornea,iris/ciliary, lens, choroid/retina and sclera. The pharmaceuticallyacceptable ophthalmic vehicle may, for example, be an ointment,vegetable oil or an encapsulating material. Alternatively, the compoundsof the invention may be injected directly into the vitreous and aqueoushumour. In a further alternative, the compounds may be administeredsystemically, such as by intravenous infusion or injection, fortreatment of the eye.

The compounds described herein may be stored in oxygen free environment.For example, a composition can be prepared in an airtight capsule fororal administration, such as Capsugel from Pfizer, Inc.

Cells, e.g., treated ex vivo with a compound as described herein, can beadministered according to methods or uses for administering a graft to asubject, which may be accompanied, e.g., by administration of animmunosuppressant drug, e.g., cyclosporin A. For general principles inmedicinal formulation, the reader is referred to Cell Therapy: Stem CellTransplantation, Gene Therapy, and Cellular Immunotherapy, by G. Morstyn& W. Sheridan eds, Cambridge University Press, 1996; and HematopoieticStem Cell Therapy, E. D. Ball, J. Lister & P. Law, ChurchillLivingstone, 2000.

Toxicity and therapeutic efficacy of compounds can be determined bystandard pharmaceutical procedures in cell cultures or experimentalanimals. The LD₅₀ is the dose lethal to 50% of the population. The ED₅₀is the dose therapeutically effective in 50% of the population. The doseratio between toxic and therapeutic effects (LD₅₀/ED₅₀) is thetherapeutic index. Compounds that exhibit large therapeutic indexes arepreferred. While compounds that exhibit toxic side effects may be used,care should be taken to design a delivery system that targets suchcompounds to the site of affected tissue in order to minimize potentialdamage to uninfected cells and, thereby, reduce side effects.

The data obtained from the cell culture assays and animal studies can beused in formulating a range of dosage for use in humans. The dosage ofsuch compounds may lie within a range of circulating concentrations thatinclude the ED₅₀ with little or no toxicity. The dosage may vary withinthis range depending upon the dosage form employed and the route ofadministration utilized. For any compound, the therapeutically effectivedose can be estimated initially from cell culture assays. A dose may beformulated in animal models to achieve a circulating plasmaconcentration range that includes the IC₅₀ (i.e., the concentration ofthe test compound that achieves a half-maximal inhibition of symptoms)as determined in cell culture. Such information can be used to moreaccurately determine useful doses in humans. Levels in plasma may bemeasured, for example, by high performance liquid chromatography.

Kits

Also provided herein are kits, e.g., kits for therapeutic purposes orkits for modulating the lifespan of cells or modulating apoptosis. A kitmay comprise one or more compounds as described herein, e.g., inpremeasured doses. A kit may optionally comprise devices for contactingcells with the compounds and instructions for use. Devices includesyringes, stents and other devices for introducing a compound into asubject (e.g., the blood vessel of a subject) or applying it to the skinof a subject.

In yet another embodiment, the invention provides a composition ofmatter comprising a compound of this invention and another therapeuticagent (the same ones used in combination therapies and combinationcompositions) in separate dosage forms, but associated with one another.The term “associated with one another” as used herein means that theseparate dosage forms are packaged together or otherwise attached to oneanother such that it is readily apparent that the separate dosage formsare intended to be sold and administered as part of the same regimen.The compound and the other agent are preferably packaged together in ablister pack or other multi-chamber package, or as connected, separatelysealed containers (such as foil pouches or the like) that can beseparated by the user (e.g., by tearing on score lines between the twocontainers).

In still another embodiment, the invention provides a kit comprising inseparate vessels, a) a compound of this invention; and b) anothertherapeutic agent such as those described elsewhere in thespecification.

The practice of the present methods will employ, unless otherwiseindicated, conventional techniques of cell biology, cell culture,molecular biology, transgenic biology, microbiology, recombinant DNA,and immunology, which are within the skill of the art. Such techniquesare explained fully in the literature. See, for example, MolecularCloning A Laboratory Manual, 2^(nd) Ed., ed. by Sambrook, Fritsch andManiatis (Cold Spring Harbor Laboratory Press: 1989); DNA Cloning,Volumes I and II (D. N. Glover ed., 1985); Oligonucleotide Synthesis (M.J. Gait ed., 1984); Mullis et al. U.S. Pat. No. 4,683,195; Nucleic AcidHybridization (B. D Hames & S. J. Higgins eds. 1984); Transcription AndTranslation (B. D. Hames & S. J. Higgins eds. 1984); Culture Of AnimalCells (R. I. Freshney, Alan R. Liss, Inc., 1987); Immobilized Cells AndEnzymes (IRL Press, 1986); B. Perbal, A Practical Guide To MolecularCloning (1984); the treatise, Methods In Enzymology (Academic Press,Inc., N.Y.); Gene Transfer Vectors For Mammalian Cells (J. H. Miller andM. P. Calos eds., 1987, Cold Spring Harbor Laboratory); Methods InEnzymology, Vols. 154 and 155 (Wu et al. eds.), Immunochemical MethodsIn Cell And Molecular Biology (Mayer and Walker, eds., Academic Press,London, 1987); Handbook Of Experimental Immunology, Volumes I-IV (D. M.Weir and C. C. Blackwell, eds., 1986); Manipulating the Mouse Embryo,(Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1986).The Examples set forth below are illustrative of the present inventionand are not intended to limit, in any way, the scope of the presentinvention.

The Examples set forth below are illustrative of the present inventionand are not intended to limit, in any way, the scope of the presentinvention.

Examples

The Examples set forth below are illustrative of the present inventionand are not intended to limit, in any way, the scope of the presentinvention, but rather to provide guidance to the skilled artisan toprepare and use the compounds, compositions, and methods or uses of thepresent invention. While particular embodiments of the present inventionare described, the skilled artisan will appreciate that various changesand modifications can be made without departing from the spirit andscope of the invention.

As used herein the symbols and conventions used in these processes,schemes and examples are consistent with those used in the contemporaryscientific literature, for example, the Journal of the American ChemicalSociety or the Journal of Biological Chemistry. Standard single-letteror three-letter abbreviations are generally used to designate amino acidresidues, which are assumed to be in the L-configuration unlessotherwise noted. Unless otherwise noted, all starting materials wereobtained from commercial suppliers and used without furtherpurification.

All references to ether are to diethyl ether; brine refers to asaturated aqueous solution of NaCl. Unless otherwise indicated, alltemperatures are expressed in ° C. (degrees Centigrade). All reactionsare conducted under an inert atmosphere at room temperature unlessotherwise noted, and all solvents are highest available purity unlessotherwise indicated.

Instrumentation Used

LCMS with PDA:

Waters Alliance 2695-2996/Quattromicro Agilent-1200/SQD

Preparative LC with UV Detector (Prep HPLC):

Waters-2545/2998 PDA and 2487 UV Shimadzu—LC-20AP/20AV-UVGilson-333,334/115-UV Chiral HPLC: Waters Alliance-2695/2998 &2996 SFCPurification Systems: Thar—SFC-80 Waters SFC—200 NMR (400 MHz):Varian-400 MHz

¹H-NMR tabulation was generated with 2014 ACD labs software.

¹H NMR (hereinafter also “NMR”) spectra were recorded on a Varian-400MHz spectromitor. Chemical shifts are expressed in parts per million(ppm, δ units). Coupling constants are in units of hertz (Hz). Splittingpatterns describe apparent multiplicities and are designated as s(singlet), d (doublet), t (triplet), q (quartet), quint (quintet), m(multiplet), br (broad).

LCMS Methods Used

Acq. Method Conditions: RND-ABC-6-MINColumn: XBridge BEH C18 (50 mm×4.6 mm, 2.5 μm)Mobile Phase: A: 5 mM Ammonium Bicarbonate in water (PH-10 withAmmonia): ACNTime (min)/% ACN: 0/5, 0.5/5, 1/15, 3.3/98, 5.2/98, 5.5/5, 6.0/5Column temp: 35° C., Flow Rate 1.3 ml/min

MS Parameters: Mass Range: 100-1000 Scan Time: 0.5 Sec

Inter-Scan delay: 0.1 sec

Run Time: 6.0 min Acq.Method Conditions: RND-FA-4.5-MIN

Column: Acquity BEH C18 (50 mm×2.1 mm, 1.7 um)Mobile Phase: A: 0.1% FA in water; B: 0.1% FA in ACNTime (min)/% B: 0/3, 0.4/3, 3.2/98, 3.8/98, 4.2/3, 4.5/3Column Temp: 35° C., Flow Rate: 0.6 mL/min

MS Parameters: Mass Range: 100-1000 Scan Time: 0.5 Sec

Inter-Scan delay: 0.1 sec

Run Time: 4.5 min Acq.Method Conditions: RND-FA-4.5-MIN

Column: Acquity BEH C18 (50 mm×2.1 mm, 1.7 um)Mobile Phase: A: 0.1% FA in water; B: 0.1% FA in ACNTime (min)/% B: 0/3, 0.4/3, 3.2/98, 3.8/98, 4.2/3, 4.5/3Column Temp: 35° C., Flow Rate: 0.6 mL/min

MS Parameters: Mass Range: 100-1000 Fragmentor: 100 Step Size: 0.1 RunTime: 4.5 min

Acq. Method Conditions: RND-ABC-6.5-MINColumn:)(Bridge BEH C18 (50 mm×4.6 mm, 2.5 μm)Mobile Phase: A: 5 mM Ammonium Bicarbonate in water (PH-10 withAmmonia): ACNTime (min)/% ACN: 0/5, 0.5/5, 1/15, 3.3/98, 6.0/98, 6.1/5, 6.5/5Column temp: 35° C., Flow Rate 1.3 ml/min

MS Parameters: Mass Range: 100-1000 Fragmentor: 100 Step Size: 0.1 RunTime: 6.5 min

Acq. Method Conditions: RND-ABC-10-MINColumn: XBridge BEH C18 (50 mm×4.6 mm, 2.5 μm)Mobile Phase: A: 5 mM Ammonium Bicarbonate in water (PH-10 withAmmonia): ACNTime (min)/% ACN: 0/5, 0.5/5, 1.5/15, 7/98, 9.0/98, 9.5/5, 10/5Column temp: 35° C., Flow Rate 1.3 ml/min

MS Parameters: Mass Range: 100-1000 Fragmentor: 100 Step Size: 0.1 RunTime: 10.0 min Intermediates Synthesis of (S)-dimethyl2-((6-chloro-3-nitropyridin-2-yl)amino)pentanedioate

To a mixture of 40.0 g (207 mmol) of 2,6-dichloro-3-nitropyridine, 87.7g (414 mmol) of L-glutamic acid dimethyl ester hydrochloride, and 69.6 g(829 mmol) of NaHCO₃ was added 600 mL of tetrahydrofuran. The mixturewas stirred at 40° C. for 24 h, monitoring for the disappearance of2,6-dichloro-3-nitropyridine by HPLC. After the reaction was complete,the solids were filtered away and washed with ethyl acetate (3×100 mL).The combined filtrate and washings were concentrated in vacuo, then theresidue was purified via silica gel chromatography, eluting with 10/1(v/v) hexanes/ethyl acetate, to give (60 g, 87%) of the product as ayellow solid, LCMS (m/z) 332.1 [M+H]⁺.

Synthesis of (S)-methyl3-(6-chloro-2-oxo-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propanoate

To a mixture of 20 g (60.2 mmol) of (S)-dimethyl2-((6-chloro-3-nitropyridin-2-yl)amino)pentanedioate and 16.8 g (301mmol) of iron powder was added 375 mL of 2-propanol, then 125 mL ofwater. To the stirred mixture was added 5.5 g (90.3 mmol) of aceticacid, then the reaction was stirred at reflux for 1 h. The reaction wasmonitored for the disappearance of starting material by HPLC. After thereaction was complete, the solids were filtered off and washed with2-propanol (3×50 mL). The combined filtrate and washings wereconcentrated to dryness, and then the residue was dried in vacuo to give15 g (81%) of the product as a dark yellow solid. This was used withoutfurther purification in the next step, LCMS (m/z) 270.1 [M+H]⁺.

Synthesis of(S)-3-(6-chloro-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propan-1-ol

To a solution of 17.78 g (133.3 mmol) of AlCl₃ in 260 mL oftetrahydrofuran (THF) under N₂ was added 200 mL of 2M LiAlH₄ in THF,dropwise, at a rate to control gas evolution. This gave a solution ofalane (AlH₃) in THF. In a separate flask, a solution of 26.0 g (96.4mmol) of (S)-methyl3-(6-chloro-2-oxo-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propanoatein 460 mL of THF was prepared under N₂, then cooled with a dryice/acetone bath. To this was added the alane solution, dropwise withstirring, over 2 h. When the addition was complete, the cooling bath wasremoved, and the reaction was allowed to warm to ambient temperature.After 1.5 h, LCMS analysis showed that the reaction was complete. Next,a solution of 17.6 g NaOH in 65 mL of water was added slowly to controlthe evolution of H₂. The suspension was allowed to stir for 18 h, andthen the solids were filtered away. The precipitate was washed withethyl acetate, then the filtrate and washings were concentrated invacuo. The product was purified via silica gel chromatography elutingwith CH₂Cl₂, followed by a gradient of 0 to 10% methanol in CH₂Cl₂ togive 15.21 g (69%) of a yellow-orange solid, LCMS (m/z) 228.1 [M+H]⁺.

Synthesis of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To 12 g (52.7 mmol) of(S)-3-(6-chloro-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propan-1-olwas added 160 mL of 48% (w/w) aq. HBr, then the reaction was stirred at90° C. for 18 h. The reaction was monitored by HPLC for thedisappearance of the starting alcohol. After the reaction was complete,it was cooled to ambient temperature, then 1.2 M aq. NaHCO₃ was addeduntil pH=8. The mixture was extracted with ethyl acetate (3×100 mL),then the combined organic phases were washed with brine (1×100 mL),dried over Na₂SO₄ and filtered, and the filtrate was concentrated todryness. The residue was purified via silica gel chromatography, elutingwith 2/1 (v/v) hexanes/ethyl acetate to give 6.0 g (55%) of the productas a light yellow solid, LCMS (m/z) 210.1 [M+H]⁺.

Synthesis of2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a solution of2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(3 g, 14.31 mmol),4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-2-(trifluoromethyl)pyridine(3.91 g, 14.31 mmol) and cesium carbonate (4.66 g, 14.31 mmol) in1,4-Dioxane (60 ml) and water (6 ml) at room temp and reaction massdegassed with argon for 20 min. Next, added solid palladium(II) acetate(3.21 g, 14.31 mmol) and x-phos (6.82 g, 14.31 mmol) in to the reactionmass in one charge. The reaction mixture was stirred at 105° C. for 3-4hrs. The reaction mass was filtered through celite bed and concentrated.The crude material was taken and dissolved in ethyl acetate and washedwith sodium bicarbonate solution and water. Organic phase was dried oversodium sulfate and concentrated to get. The residue was triturated withn-pentane (3×50 mL). The resulting solid was filtered through a Buchnerfunnel, rinsed with n-pentane, and collected as the desired product (4g, 86%), LCMS (m/z) 321.3 (M+H)⁺.

Synthesis of(9S)-2-(5-(trifluoromethyl)pyridin-3-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a solution of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(3 g, 14.31 mmol), (5-(trifluoromethyl)pyridin-3-yl)boronic acid (5.46g, 28.6 mmol) and Cs2CO3 (13.99 g, 42.9 mmol) in Tetrahydrofuran (THF)(60 ml), water (4 ml) stirred under nitrogen at 25° C., purged withArgon gas for 20 minutes. Then palladium (II) acetate (0.080 g, 0.358mmol) and X-Phos (227 mg) were added. The reaction mixture was stirredat 110° C. for 16 hr. Next, the reaction mixture was concentrated andthe residue was taken up in DCM (100 mL). The solution was washed withwater and brine, dried over Na₂SO₄, filtered and concentrated. The crudeproduct was added to a silica gel column and was eluted with Hex/EtOAc(1:1) Collected fractions were evaporated to give as a off white solid(3.6 g, 11.1 mmol 76%), LCMS (m/z) 321.2 [M+H]⁺.

Synthesis of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a solution of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(10 g, 47.7 mmol), (2-methylpyridin-4-yl)boronic acid (8.49 g, 62.0mmol) and Potassium phosphate (30.4 g, 143 mmol) in 1-Butanol (300 ml),water (100 ml) stirred under nitrogen at 25° C., purged with Argon gasfor 20 minutes was added X-Phos (2.274 g, 4.77 mmol), Pd2(dba)3 (2.184g, 2.385 mmol). The reaction mixture was stirred at 120° C. for 16 hr.Before being concentrated and the residue was taken up in DCM (700 mL).The solution was washed with water and brine, dried over Na₂SO₄,filtered and concentrated to get brown semisolid crude product. Thecrude product was purified by washing with hexane to get off white solidproduct (9 g, 32.4 mmol, 67.9% yield), LCMS (m/z): 267.3 (M+H)⁺.

Synthesis of(9S)-2-((R)-2-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a suspension of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 2.385 mmol), (R)-2-(trifluoromethyl)pyrrolidine (0.663 g, 4.77mmol) and KO^(t)Bu (0.535 g, 4.77 mmol) in 1,2-Dimethoxyethane (DME) (10mL) under nitrogen atmosphere at room temperature was added solid[1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene]chloro][3-phenylallyl]palladium(II)(1.549 g, 2.385 mmol) and stirred at 80° C. for 16 h. The reaction masswas cooled down to room temperature and filtered through celite and thesolvent was evaporated under reduced pressure to obtain crude residue.The crude residue was diluted with EtOAc (100 mL) and washed with water(50 mL×2) followed by brine solution and dried over anhydrous sodiumsulfate, filtered and evaporated to obtain the crude product. The crudeproduct was purified by flash column chromatography (silica gel: 100-200mesh, eluted with 1:1 Hex/EtOAc) to afford(9S)-2-((2R)-2-(trifluoromethyl)cyclopentyl)-6,7,8,9,10,10a-hexahydro-4aH-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.596 mmol, 64.6% yield) as a light green solid (TLC: R_(f)0.3, eluent: 80% EtOAc in Hexane), LCMS (m/z) 313.2 [M+H]⁺.

Synthesis of(9S)-2-((S)-2-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

A suspension of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 2.385 mmol), (S)-2-(trifluoromethyl)pyrrolidine (0.663 g, 4.77mmol) and KOtBu (0.535 g, 4.77 mmol) in 1,2-Dimethoxyethane (DME) (10mL) was stirred and degassed with argon at room temp for 15 mins. Next,[1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene]chloro][3-phenylallyl]palladium(II)(1.549 g, 2.385 mmol) added to the reaction mixture. Then the reactionmixture was stirred 16 hr at 80° C. After completion, the reaction massfiltered through celite and concentrated to dryness. The resultingresidue was diluted with EtOAc and washed with water followed by brinesolution and dried out with sodium sulfate, filtered and evaporated. Thecrude product was added to a silica gel column, eluted with Hex/EtOAc(1:1). Collected fractions were evaporated to get (9S)-2-((S)-2-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.6 g, 1.868 mmol, 78% yield) as an off-white solid, LCMS (m/z) 313.3[M+H]⁺.

Synthesis of(9S)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

KOtBu (0.321 g, 2.86 mmol was added to a stirred solution of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.3 g, 1.431 mmol), and 3-(trifluoromethyl)pyrrolidine (0.398 g, 2.86mmol) in 1,2-Dimethoxyethane (DME) (10 mL). The reaction mixture wasstirred and degassed with argon at room temp for 15 mins. and then(1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene)chloro)(3-phenylallyl)palladium(2) (0.037 g, 0.057 mmol) added to the reactionmixture. The reaction was stirred for 16 h at 80° C. The reaction wascooled to room temperature, filtered through celite and evaporated underreduced pressure. The reaction mixture was partitioned between water (20mL) and EtOAc (50 mL). Organic layer was separated and was dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude asbrown solid (TLC eluent: 80% EtOAc: Rf-0.4; UV active). The crude waspurified by column chromatography using (100-200 mesh) silica gel andwas eluted with 50% EtOAc in Hexane to afford (85:15) mixture andfurther purified by SFC to afford pure(9S)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.151 g, 0.473 mmol, 33.1% yield) as a Off-white solid.

Analytical SFC Conditions: Peak-II Column/dimensions: Chiralpak AD-H(250×4.6) mm, 5 u % CO2: 70.0%

% Co solvent: 30.0% (0.5% DEA In MeOH)Total Flow: 3.0 g/min

Back Pressure: 100 bar Temperature: 26.8° C. UV: 212 nm Preparative SFCConditions Column/dimensions: Chiralpak AD-H (250×30) mm % CO2: 75%

% Co solvent: 25.0% (MeOH)Total Flow: 100.0 g/min

Back Pressure: 100 bar UV: 212 nm

Stack time: 1.8 min

Load/inj: 5.5 mg

LCMS (m/z) 313.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d6): δ 6.87 (d, J=8.1 Hz, 1H), 6.49 (d, J=4.9 Hz,1H), 5.58 (d, J=8.1 Hz, 1H), 3.60 (dd, J=10.8, 8.4 Hz, 1H), 3.47 (s,1H), 3.39 (dd, J=10.6, 5.8 Hz, 2H), 3.30 (s, 2H), 3.14-2.98 (m, 2H),2.90 (d, J=4.5 Hz, 1H), 2.50 (qd, J=3.2, 2.2, 1.6 Hz, 1H), 2.31-2.13 (m,1H), 2.05 (d, J=7.1 Hz, 1H), 1.72 (dt, J=7.0, 3.0 Hz, 2H), 1.44 (s, 1H),1.16 (d, J=14.4 Hz, 1H).

Synthesis of(9S)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

(1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene)chloro)(3-phenylallyl) palladium (0.062 g, 0.095 mmol) added to a degassedsuspension of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 2.385 mmol), 3-(trifluo-romethyl)pyrrolidine (0.663 g, 4.77mmol) and KOtBu (0.535 g, 4.77 mmol) in 1,2-dimethoxyethane (20 mL) atRT. The reaction mixture was further degassed for 10 min and was stirredfor 16 h at 80° C. The reaction mixture was cooled to RT and wasfiltered through a pad of celite. The filtrate was evaporated to obtainbrown residue. The residue was partitioned between water (15 mL) andEtOAc (2×25 mL). The organic layer was washed with water followed bybrine solution and dried over anhydrous sodium sulfate, filtered andfiltrate was evaporated to get the crude (TLC eluent: 80% EtOAc/hexane,Rf value: 0.4, UV active). The crude was purified by columnchromatography (100-200 mesh) using silica gel, and the product waseluted 50% ethyl acetate in pet ether to give (30:70) mixture ofenantiomers which on further SFC purification afforded(9S)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diaz-ocine(0.125 g, 0.393 mmol, 16.46% yield) as a brown solid.

Analytical SFC Conditions: Peak I (major)

Chiralpak AD-H (250×4.6) mm, 5 u % CO2: 70.0%

% Co solvent: 30.0% (0.5% DEA In MeOH)Total Flow: 3.0 g/min

Back Pressure: 100 bar Temperature: 26.8° C. UV: 212 nm Preparative SFCConditions Column/dimensions: Chiralpak AD-H (250×30) mm % CO2: 75%

% Co solvent: 25.0% (MeOH)Total Flow: 100.0 g/min

Back Pressure: 100 bar UV: 212 nm

Stack time: 1.8 min

Load/inj: 5.5 mg

LCMS (m/z) 313.31[M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 7.03 (dd, J=8.2, 0.6 Hz, 1H), 5.67 (d, J=8.2Hz, 1H), 4.91 (s, 1H), 3.71 (dd, J=10.8, 8.5 Hz, 1H), 3.62 (s, 1H), 3.52(dd, J=10.8, 7.2 Hz, 2H), 3.43 (dt, J=9.6, 7.5 Hz, 1H), 3.24-3.08 (m,3H), 2.99 (d, J=8.2 Hz, 1H), 2.83 (d, J=12.0 Hz, 1H), 2.27-2.08 (m, 2H),1.91-1.75 (m, 2H), 1.65 (d, J=5.0 Hz, 2H), 1.26 (t, J=6.6 Hz, 1H).

Synthesis of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

A suspension of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.750 g, 3.58 mmol), 3-(trifluoromethyl)piperidine (1.096 g, 7.15 mmol)and KOtBu (1.204 g, 10.73 mmol) in 1,2-Dimethoxyethane (DME) (30 mL)stirred and degassed with argon at room temp for 15 mins. and then(1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene)chloro)(3-phenylallyl)palladium(2) (0.093 g, 0.143 mmol) added to the reactionmixture. Then the reaction mixture was stirred for 16 h at 80° C. Thereaction mass filtered through celite and evaporated under reducedpressure completely. Reaction mixture was diluted with EtOAc (50 ml),washed with water followed by brine solution, dried with sodium sulfate,filtered and evaporated. The crude product was added to a silica gelcolumn and was eluted with Hex/EtOAc (1:1). Collected fractions wereevaporated and the resulting residue was purified via chiral SFCseparation to get pure(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.450 g, 1.371 mmol, 38.3% yield) as off white solid, (TLC: R_(f)value: 0.4, 80% EtOAc/Hexane).

Analytical SFC Conditions: Peak-I Column/dimensions: Chiralpak AD-H(250×4.6) mm, 5 u % CO2: 60.0%

% Co solvent: 40.0% (0.5% DEA IN MeOH)Total Flow: 4.0 g/min

Back Pressure: 100 bar Temperature: 26.8° C. UV: 215 nm Preparative SFCConditions Column/dimensions: Chiralpak AD-H (250×30) mm % CO2:65.0%

% Co solvent: 35.0% (100% MeOH)Total Flow: 100.0 g/min

Back Pressure: 100 bar UV: 215 nm

Stack time: 2.2 min

Load/inj: 50.0 mg

LCMS (m/z) 327.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ 6.89 (dd, J=8.2, 0.7 Hz, 1H), 6.56 (d,J=4.6 Hz, 1H), 5.89 (d, J=8.3 Hz, 1H), 4.58-4.33 (m, 1H), 3.97 (d,J=12.8 Hz, 1H), 3.48 (s, 1H), 3.17-2.98 (m, 2H), 2.92 (d, J=4.4 Hz, 1H),2.76-2.57 (m, 2H), 2.50 (p, J=1.9 Hz, 2H), 1.96 (d, J=10.6 Hz, 1H), 1.74(dd, J=9.4, 3.3 Hz, 3H), 1.61-1.32 (m, 3H), 1.25-1.08 (m, 1H).

Synthesis of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

KOtBu (0.803 g, 7.15 mmol) was added to a stirred solution of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.750 g, 3.58 mmol), and 3-(trifluoromethyl)piperidine (1.096 g, 7.15mmol) in 1,2-Dimethoxyethane (DME) (20 mL). The reaction was stirred anddegassed with argon at room temp for 15 mins. and then(1,3-Bis(2,6-di-isopropylphenyl)-4,5-dihydroimidazol-2-ylidene)chloro)(3-phenylallyl)palladium(2) (0.093 g, 0.143 mmol) added to the reactionmixture. The reaction mixture was stirred for 16 h at 80° C. Thereaction was cooled to room temperature, filtered through celite andevaporated completely, and was partitioned between water (20 mL) andEtOAc (50 mL). Organic layer was separated and was dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to give crude as brownsolid (TLC eluent: 80% EtOAc: Rf-0.4; UV active). The crude was purifiedby column chromatography using (100-200 mesh) silica gel and was elutedwith 50% EtOAc in Hexane to afford (69: 31) mixture and further purifiedby SFC to afford pure(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.310 mmol, 36.6% yield) as a pale yellow solid.

Analytical SFC Conditions: Peak-II Column/dimensions: Chiralpak AD-H(250×4.6) mm, 5 u % CO2: 60.0%

% Co solvent: 40.0% (0.5% DEA IN MeOH)Total Flow: 4.0 g/min

Back Pressure: 100 bar Temperature: 26.8° C. UV: 215 nm Preparative SFCConditions Column/dimensions: Chiralpak AD-H (250×30) mm % CO2: 65.0%

Co solvent: 35.0% (100% MeOH)Total Flow: 100.0 g/min

Back Pressure: 100 bar UV: 215 nm

Stack time: 2.2 min

Load/inj: 50.0 mg

LCMS (m/z) 327.2 [M+H]⁺.

1H NMR (400 MHz, DMSO-d6): δ ppm 6.89 (d, J=8.11 Hz, 1H), 6.55 (br d,J=3.95 Hz, 1H), 5.89 (d, J=8.33 Hz, 1H), 4.49 (dt, J=12.44, 1.78 Hz,1H), 3.96 (d, J=13.15 Hz, 1H), 3.48 (br s, 1H), 3.15-2.96 (m, 2H),2.94-2.80 (m, 1H), 2.74-2.56 (m, 2H), 2.55-2.41 (m, 2H), 2.12-1.84 (m,1H), 1.82-1.64 (m, 3H), 1.58-1.32 (m, 3H), 1.32-1.08 (m, 1H).

Synthesis of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a solution of(9S)-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(2 g, 6.26 mmol) in Chloroform (20 mL) stirred under nitrogen at 0° C.was added NCS (1.004 g, 7.52 mmol). The reaction mixture was stirred atRT for 2 hr. Reaction mixture was quenched with water and extracted with2×25 ml of DCM, organic layer was dried over Na₂SO₄ and concentratedunder reduced pressure to afford crude compound. The crude product waspurified by flash column chromatography with 100-200 silica gel and waseluted with 70% ethyl acetate in pet ether to afford pure compound(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(800 mg, 1.859 mmol, 29.7% yield) as pale yellow semi solid, LCMS (m/z):354.22 [M+H]⁺.

Synthesis of(9S)-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To solid(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(5 g, 23.85 mmol), (3-(trifluoromethyl)phenyl)boronic acid (6.79 g, 35.8mmol) and potassium phosphate tri basic (15.19 g, 71.5 mmol) in1,4-Dioxane (160 mL) and Water (40 mL) stirred and degas with argon for10 mints then added solid x-phos (0.689 g, 4.77 mmol) and Pd₂(dba)₃(2.184 g, 2.385 mmol) again the reaction mixture degas for 5 mints andstirred at 110° C. for 16 hr. The organic phase was washed with water 50mL, saturated brine 100 mL and dried over Na₂SO₄ and evaporated undervacuum to give the crude products as a brown solid. The crude productswas washed with diethyl ether and pentane and filtered washed withdiethyl ether to get pure compound(9S)-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(4.8 g, 13.81 mmol, 57.9% yield), LCMS (m/z): 320.11 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (377 mg, 1.272 mmol) was added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.272 mmol), and TEA (0.886 mL, 6.36 mmol) in Tetrahydrofuran(THF) (50 mL) at 28° C. The reaction mixture was stirred for 30 min andwas added (R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine(571 mg, 2.54 mmol). The reaction mixture was stirred for 10 h at 70° C.The reaction mixture was cooled to room temperature and was partitionedbetween water (5 mL) and EtOAc (15 mL). EtOAc layer was separated andwas dried over anhydrous Na₂SO₄, filtered. The filtrate was evaporatedto get crude. The crude was purified by GRACE using C-18 reservalcolumn, Mobile phase A: 0.1% Formic Acid in water; B: MeOH, the productwas eluted at 91% of MeOH in 0.1% Formic Acid in water. The solvent wasevaporated and was basified with saturated NaHCO₃. The aqueous layer wasextracted with DCM, DCM layer was dried over anhydrous Na₂SO₄, filteredand filtrate was evaporated to afford(9S)-3-chloro-N-(6-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.391 mmol, 30.8% yield) as yellow solid, LCMS (m/z): 604.14[M+H]⁺.

Synthesis of(9S)-2-chloro-3-iodo-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

NIS (2.58 g, 11.45 mmol) was added to a stirred solution of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(2.0 g, 9.54 mmol) in Chloroform (40 mL) under nitrogen at 0° C. and thereaction mixture was stirred 16 hr at 65° C. The reaction mixture wascooled to room temp, solvent evaporated under reduced pressurecompletely to afford the crude product. The crude product was purifiedby column chromatography using neutral alumina and was eluted with 20%EtOAc in Hexane (gradient system) to afford the desired product(9S)-2-chloro-3-iodo-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1.65 g, 4.53 mmol, 47.5% yield) as a pale yellow solid, LCMS (m/z):335.90 [M+H]⁺.

Synthesis of(9S)-2-chloro-3-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

A suspension of(9S)-2-chloro-3-iodo-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1.65 g, 4.92 mmol), trimethylboroxine (1.639 mL, 4.92 mmol) andpotassium carbonate (2.039 g, 14.75 mmol) in 1,4-Dioxane (15 mL) & Water(1.5 mL) stirred and degassed with argon at room temp for 15 mins,tetrakis(triphenylphosphine)palladium(0) (0.568 g, 0.492 mmol) was addedto the reaction mixture. Then the reaction mixture was stirred 48 hr at90° C. The reaction mixture was cooled to room temp, and filteredthrough celite and washed with EtOAc (30 ml). Take filtrate andconcentrated and dissolved with EtOAc (50 ml). EtOAc layer washed withwater (15 ml) followed by brine solution (15 ml) and dried out withNa₂SO₄, filtered and concentrated to get crude product. The crudeproduct was purified by column chromatography using neutral alumina andwas eluted with 20% EtOAc in Hexane (gradient system) to afford thedesired product(9S)-2-chloro-3-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.800 g, 3.55 mmol, 72.1% yield) as a pale yellow solid (TLC eluent:50% EtOAc in Hexane: R_(f)-0.3; UV active). LCMS (m/z): 224.90 [M+H]⁺.

Synthesis of(9S)-3-methyl-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

A suspension of(9S)-2-chloro-3-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(800 mg, 3.58 mmol), (2-methylpyridin-4-yl)boronic acid (612 mg, 4.47mmol) and tripotassium phosphate (2277 mg, 10.73 mmol) in 1,4-Dioxane(20 mL) & Water (4 mL) stirred and degassed with argon at room temp for15 mins. Pd₂(dba)₃ (164 mg, 0.179 mmol) and X-Phos (170 mg, 0.358 mmol)added to the reaction mixture. Then the reaction mixture was stirred 16hr at 90° C. The reaction was monitored by TLC. The reaction mixture wascooled to room temp and filtered through celite and washed with EtOAc.Take filtrate and concentrated and dissolved with EtOAc. EtOAc layerwashed with water followed by brine solution and dried out with Na₂SO₄,filtered and concentrated to get crude product. The crude product waspurified by ether (20 ml) washings to afford desired product(9S)-3-methyl-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(600 mg, 2.121 mmol, 59.3% yield) as an off white solid, LCMS (m/z):281.16 [M+H]⁺.

Synthesis of 2,6-dichloro-4-methyl-3-nitropyridine

To a suspension of 2,6-dichloro-4-methylpyridine (5 g, 30.9 mmol) intrifluoroacetic anhydride (24.98 mL, 177 mmol) cooled to 0° C. was addeddropwise nitric acid (2.90 mL, 64.8 mmol) into it. The resultingsolution was stirred at RT for 18 hr. The reaction mixture was addedslowly to a chilled solution of sodium metabisulfite (5.87 g, 30.9 mmol)in Water (40 mL) and stirred at RT for 2 hr. The reaction mixture wasneutralized to pH 7 using 8N NaOH (25 mL) solution and extracted twicewith DCM (100 mL). The combined organic layers were washed with brine,dried over Na₂SO₄, filtered and concentrated under reduced pressure toafford 2,6-dichloro-4-methyl-3-nitropyridine (6.2 g, 29.4 mmol, 95%yield) as off white solid, LCMS (m/z): 206.98 [M+H]⁺.

Synthesis of (S)-dimethyl2-((6-chloro-4-methyl-3-nitropyridin-2-yl)amino)pentanedioate

Procedure: (S)-dimethyl 2-aminopentanedioate hydrochloride (147 g, 696mmol) was added to a suspension of 2,6-dichloro-4-methyl-3-nitropyridine(120 g, 580 mmol) and sodium bicarbonate (146 g, 1739 mmol) inTetrahydrofuran (THF) (2.5 L) at 0° C. under nitrogen. The reactionmixture was stirred at 65° C. for 24 hr. The reaction was monitored byTLC. The reaction mixture was filtered and washed with EtOAc (2×30 mL).The filtrate was concentrated under vacuum to afford crude. The crudewas purified by column chromatography using silica gel (100-200 mesh),and the product was eluted with 10% EtOAc in Pet ether to afford(S)-dimethyl2-((6-chloro-4-methyl-3-nitropyridin-2-yl)amino)pentanedioate (50 g, 135mmol, 23.31% yield) as yellow solid, LCMS (m/z): 345.1 [M+H]⁺.

Synthesis of (S)-methyl3-(6-chloro-8-methyl-2-oxo-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propanoate

Iron (40.4 g, 723 mmol) was added to a stirred solution of (S)-dimethyl2-((6-chloro-4-methyl-3-nitropyridin-2-yl)amino)pentanedioate (50 g, 145mmol) in Isopropanol (450 mL) and Water (90 mL) at room temp. Thereaction mixture was heated to 40° C. and was added acetic acid (12.42mL, 217 mmol). The reaction mixture was stirred at 80° C. for 2 hr. Thereaction was monitored by TLC. The reaction mixture was cooled to RT,and basified with saturated NaHCO₃, filtered through a pad of celite andwas washed with DCM (3×50 mL). The filtrate was separated and was washedwith brine solution, dried over anhydrous Na₂SO₄, filtered and filtratewas concentrated under reduced pressure to afford crude product. Thecrude product was purified by washed with ether (20 ml) to afforddesired product (S)-methyl3-(6-chloro-8-methyl-2-oxo-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propanoate(29 g, 101 mmol, 69.8% yield) as an off-white solid, LCMS (m/z): 284.06[M+H]⁺.

Synthesis of(S)-3-(6-chloro-8-methyl-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propan-1-ol

Procedure: 2M lithium aluminum hydride (256 mL, 511 mmol) was addeddropwise to a stirred solution of aluminum chloride (19.08 g, 143 mmol),in Tetrahydrofuran (THF) (290 mL) under nitrogen at a rate to controlgas evolution. The reaction mixture was stirred for 30 min at rt. Thereaction mixture (alane AlH₃) was added dropwise to a stirred solutionof (S)-methyl3-(6-chloro-8-methyl-2-oxo-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propanoate(29 g, 102 mmol) in Tetrahydrofuran (THF) (450 mL) dropwise at −78° C.under nitrogen over 30 minutes. The reaction was allowed to warm toambient temperature for 16 hr. The reaction was monitored by TLC. Thereaction mixture was quenched with 10% NaOH solution at 0° C. andstirred 16 hr and filtered through a pad of celite and was washed with(300 ml) DCM. DCM layer was dried over anhydrous Na₂SO₄, filtered andfiltrate was evaporated to afford the crude product. The crude productwas purified washing with ether (2×50 ml) to afford(S)-3-(6-chloro-8-methyl-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propan-1-ol(22 g, 69.8 mmol, 68.3% yield) as an off-white yellow solid, LCMS (m/z):242.0 [M+H]⁺.

Synthesis of(9S)-2-chloro-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a stirred solution of(S)-3-(6-chloro-8-methyl-1,2,3,4-tetrahydropyrido[2,3-b]pyrazin-3-yl)propan-1-ol(21 g, 87 mmol) was added HBr (11.79 ml, 217 mmol) at 28° C. and stirredfor 12 hr at 100° C. The reaction was monitored by TLC. The reactionmixture was cooled to 28° C., and was poured in to ice water (50 mL).The aqueous layer was neutralized with saturated NaHCO₃ solution (120ml). The aqueous layer was extracted with EtOAc (2×50 ml). EtOAc layerwas dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated tothe crude. The crude was purified by column chromatography using neutralalumina and the product was eluted with 50% EtOAc in Pet ether to affordthe compound. The compound was washed with n-pentane to afford pure(9S)-2-chloro-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(13 g, 57.8 mmol, 66.5% yield) as white solid, LCMS (m/z): 224.09[M+H]⁺.

Synthesis of(9S)-4-methyl-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a degassed solution of(9S)-2-chloro-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 2.235 mmol), (2-methylpyridin-4-yl)boronic acid (459 mg, 3.35mmol) and K₃PO₄ (1423 mg, 6.71 mmol) in 1,4-Dioxane (40 mL); Water (10mL) and was added x-phos (213 mg, 0.447 mmol), Pd₂(dba)₃ (205 mg, 0.224mmol). The reaction mixture was stirred at 110° C. for 12 hr. Thereaction mixture was cooled to 28° C. and was partitioned between water(15 mL) and EtOAc (2×15 mL). EtOAc layer was separated and was driedover anhydrous Na₂SO₄, filtered. The filtrate was evaporated to getcrude. The crude was purified by washing with diethyl ether (15 mL) toafford pure(9S)-4-methyl-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(550 mg, 1.920 mmol, 86% yield) as yellow solid, LCMS (m/z): 281.05[M+H]⁺.

Synthesis of(9S)-3-chloro-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine

To a solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(2.0 g, 7.51 mmol) in Chloroform (20 mL) stirred under nitrogen at 0° C.was added NCS (1.504 g, 11.26 mmol) portionwise during 10 min. Thereaction mixture was stirred at 0° C. for 4 hr. Reaction mass wasquinched with the ice cold water (20 ml) and extracted with the DCM(3×30 ml). Organic phase was separated and washed with the water (2×20ml) and brine solution (2×20 ml), then organic phase was dried overanhydrous sodium sulphate, filtered it and filterate was evaporatedunder reduced pressure to get the crude product. The crude product(N35964-5-A1 & N35964-7-A1) was purified by column chromatography usingneutral alumina and was eluted with 50% EtOAc in Hexane (gradientsystem) to afford the desired product(9S)-3-chloro-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1.2 g, 3.60 mmol, 47.9% yield) as a pale yellow solid, LCMS (m/z):300.17 [M+H]⁺.

Synthesis of (9S)-phenyl3-chloro-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxylate

To a solution of phenyl carbonochloridate (0.195 mL, 1.555 mmol) andpyridine (0.149 mL, 1.837 mmol) in Dichloromethane (DCM) (35 mL) stirredunder nitrogen at 0° C. was added(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.413 mmol). The reaction mixture was stirred at RT for 2 h.The Reaction was monitored by TLC. The reaction mixture was quenchedwith saturated sodium bicarbonate solution and extracted with DCM (200mL) twice. Combined DCM layer washed with water (80 mL) and dried outwith Na₂SO₄, filtered and concentrated under high vacuum to get crudeproduct. The crude product was added to a silica gel (100-200 mesh)column and was eluted with 30% Hex/EtOAc the collected fraction wasdistilled under reduced pressure to afford a compound. The compound waswashed with pentane to get a pure compound of (9S)-phenyl3-chloro-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxylate(500 mg, 1.044 mmol, 73.9% yield) as a white solid, LCMS (m/z): 474.30[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (377 mg, 1.272 mmol) was added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.272 mmol), and TEA (0.886 mL, 6.36 mmol) in Tetrahydrofuran(THF) (50 mL) at 28° C. The reaction mixture was stirred for 30 min andwas added (S)-6-(2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine(571 mg, 2.54 mmol). The reaction mixture was stirred for 10 h at 70° C.The reaction mixture was cooled to room temperature and was partitionedbetween water (15 mL) and EtOAc (25 mL). EtOAc layer was separated andwas dried over anhydrous Na₂SO₄, filtered. The filtrate was evaporatedto get crude. The crude was purified by GRACE using C-18 reservalcolumn, Mobile phase A: 0.1% Formic Acid in water; B: MeOH, the productwas eluted at 91% of MeOH in 0.1% Formic Acid in water. The solvent wasevaporated and was basified with saturated NaHCO₃. The aqueous layer wasextracted with DCM, DCM layer was dried over anhydrous Na₂SO₄, filtered,and filtrate was evaporated to afford(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(270 mg, 0.430 mmol, 33.8% yield) as yellow solid, LCMS (m/z): 606.11[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

TEA (1.064 mL, 7.63 mmol) and triphosgene (377 mg, 1.272 mmol) was addedto a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.272 mmol) in Tetrahydrofuran (THF) (50 mL) under nitrogen atroom temp. The reaction mixture was stirred at RT for 30 min.(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (573 mg,2.54 mmol) was added and the reaction mixture was stirred 16 hr at 65°C. The reaction mixture was cooled to room temp, solvent evaporatedunder reduced pressure completely and was partitioned between water (10mL) and EtOAc (50 mL). Organic layer was separated, dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to afford crude product.The crude product was purified by column chromatography using neutralalumina and was eluted with 50% EtOAc in Hexane (gradient system) toafford the desired product(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.271 mmol, 21.27% yield) as an off white solid, LCMS (m/z):605.2 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

TEA (1.064 mL, 7.63 mmol) and triphosgene (377 mg, 1.272 mmol) was addedto a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.272 mmol) in Tetrahydrofuran (THF) (45 mL) under nitrogen atroom temp. The reaction mixture was stirred at RT for 30 min.(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-amine (860mg, 3.82 mmol) was added and the reaction mixture was stirred 16 hr at65° C. The reaction mixture was cooled to room temp, solvent evaporatedunder reduced pressure completely and was partitioned between water (30mL) and EtOAc (100 mL). Organic layer was separated, dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to afford crudeproduct. The crude product was purified by column chromatography usingneutral alumina and was eluted with 20% EtOAc in Hexane (gradientsystem) to afford the desired product(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.317 mmol, 24.91% yield) as a pale yellow solid, LCMS (m/z):605.19 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

A solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.4 g, 1.131 mmol) and TEA (0.788 mL, 5.65 mmol) in Tetrahydrofuran(THF) (50 mL) was added triphosgene (0.336 g, 1.131 mmol) and stirredunder nitrogen at room temp for 1 h. To this reaction mixture(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine (0.382g, 1.696 mmol) was added. The reaction mixture was stirred at 65° C. for16 h and progress of the reaction was monitored by TLC and LCMS. Thereaction mixture was cooled to room temperature, poured in to water (50mL) and extracted with EtOAc (2×50 mL). The organic layer was washedwith water and brine. The organic layer was filtered through Na₂SO₄ andconcentrated to obtain crude compound. The crude compound was purifiedby neutral alumina and eluted in 50% ethyl acetate in hexane to afford(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.3 g, 0.486 mmol, 43.0% yield) as off white solid, LCMS (m/z): 605.55[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (417 mg, 1.405 mmol) was added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-6,7,8,9,10,10a-hexahydro-4aH-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.405 mmol) and TEA (1.175 mL, 8.43 mmol) in Tetrahydrofuran(THF) (50 mL) at room temp. The reaction mixture was stirred for 1 h and(R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-amine (791mg, 3.51 mmol) was added. The reaction mixture was stirred at 65° C. for16 h. Reaction was monitored by TLC. The reaction mixture was evaporatedunder reduced pressure and diluted with water (50 mL) and extracted withEthyl acetate (2×75 mL) and followed by brine solution (50 mL) andseparated the layer, dried with anhydrous Na₂SO₄, filtered andconcentrated to get crude product. The crude product was submitted toNeutral Alumina column by eluting 50-60% Ethyl acetate in Hexane. Thecollected fraction was evaporated under reduced pressure to get the purecompound of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(350 mg, 0.544 mmol, 38.7% yield) as a Off white solid, LCMS (m/z):605.23 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (252 mg, 0.848 mmol) followed by triethylamine (1.182 mL,8.48 mmol) were added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.413 mmol) in Tetrahydrofuran (THF) (15 mL) at RT and stirredfor 30 min. Then(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine (637mg, 2.83 mmol) were added to the reaction mixture at RT and stirred at80° C. for 15.5 h. Reaction mixture was cooled to RT, diluted with water(40 mL), extracted with ethyl acetate (2×60 mL) and washed with brinesolution (30 mL). Organic layer was separated, dried over Na₂SO₄,filtered and concentrated to get crude compound. The crude compound waspurified by column chromatography using silica gel (100-200 mesh), 30%ethyl acetate in pet ether as an eluent to afford(9S)-3-chloro-N-(6-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.324 mmol, 22.92% yield) as brown gummy solid, LCMS (m/z):606.99 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (419 mg, 1.413 mmol) was added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.413 mmol), and TEA (1.182 mL, 8.48 mmol) in Tetrahydrofuran(THF) (50 mL) under nitrogen at 28° C. The reaction mixture was stirredat RT for 30 min. and was added(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (955mg, 4.24 mmol) The reaction mixture was stirred 16 hr at 65° C. Thereaction mixture was cooled to 28° C., the reaction mixture waspartitioned between water (20 mL) and EtOAc (2×25 mL). Organic layer wasseparated and was dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to give crude. The crude was purified by GRACE using C-18reserval column, Mobile phase A: 0.1% Formic Acid in water; B: ACN, theproduct was eluted at 50% of ACN in 0.1% Formic Acid in water. Thesolvent was evaporated and was basified with saturated NaHCO₃. Theprecipitated solid was filtered, and was dried to afford(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(120 mg, 0.191 mmol, 13.48% yield) as off white solid, LCMS (m/z):605.23 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (0.419 g, 1.413 mmol) was added to a solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 1.413 mmol), TEA (0.985 mL, 7.07 mmol)) in Tetrahydrofuran (THF)(15 mL) was stirred under nitrogen at room temp for 1 h. To thisreaction mixture(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (0.637g, 2.83 mmol) was added. The reaction mixture was stirred at 65° C. for16 h and progress of the reaction was monitored by TLC and LCMS. Thereaction mixture was cooled to room temperature, poured in to ice water(50 mL) and extracted with EtOAc (2×100 mL). The combined organic layerwas washed with water (50 mL), brine solution (50 mL), dried overNa₂SO₄, filtered and evaporated to obtain crude compound. The crudecompound was purified by column chromatography using neutral alumina andeluted in 50% EtOAc in hexane to afford(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.52 g, 0.387 mmol, 27.4% yield) as pale yellow solid, LCMS (m/z):605.20 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(600 mg, 1.696 mmol) followed by triphosgene (302 mg, 1.018 mmol) wereadded to a solution of(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (458 mg,2.035 mmol) and TEA (1.182 mL, 8.48 mmol) in Tetrahydrofuran (THF) (20mL) at 25° C., stirred for 16 h at 70° C. Progress of the reaction wasmonitored by LCMS and TLC. The reaction mixture was cooled to 28° C. andwas partitioned between water (20 mL) and EtOAc (2×50 mL). Organic layerwas separated and was dried over anhydrous Na₂SO₄, filtered and filtratewas evaporated to get crude. The sample was loaded in dichloromethaneand purified on silica 5 g using a 0-15% methanol-dichloromethane over80 min. The appropriate fractions were combined and evaporated in vacuumto give the required product 220 mg as an off-white solid, LCMS (m/z):605.17 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (637 mg,2.83 mmol) in Tetrahydrofuran (THF) (20 mL) stirred under nitrogen atroom temp was added triphosgene (419 mg, 1.413 mmol) and triethylamine(1.182 mL, 8.48 mmol), To this(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (637 mg,2.83 mmol) was added and the reaction mixture was stirred at 65° C. for16 hr. Reaction mixture was quenched with ice water and extracted with2×25 ml of ethyl acetate, combined organic layers were dried over Na₂SO₄and concentrated under reduced pressure to afford crude compound. Thecrude product was purified by flash column chromatography (100-200silica gel) eluting at 2% methanol in DCM to afford pure compound(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.308 mmol, 21.82% yield) as pale brown solid, LCMS (m/z):605.1 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1 g, 2.83 mmol) was dissolved in Tetrahydrofuran (THF) (30 mL) stirredunder nitrogen at 0° C. were added triphosgene (0.839 g, 2.83 mmol),DIPEA (2.468 mL, 14.13 mmol). The reaction mixture was stirred for 16 atRT. To this(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (1.268 g,5.65 mmol) was added and stirred for 16 h at 80° C. in sealed tube. Thereaction mixture allowed to room temperature and quenched with 200 ml ofwater and extracted with 3×200 ml of ethyl acetate, the combined organiclayer was dried over Na₂SO₄ and concentrated under reduced pressure toobtain crude compound. The crude product was purified by flash columnchromatography to afford(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.486 mmol, 17.18% yield) as an off white solid. LCMS (m/z):604.20 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

TEA (1.970 mL, 14.13 mmol) followed by triphosgene (0.839 g, 2.83 mmol)were added to a solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1 g, 2.83 mmol) in Tetrahydrofuran (THF) (20 mL) at RT and stirred for6 h at RT and(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (0.697 g,3.11 mmol) was added and heated at 80° C. for 10 h. The reaction mixturewas cooled to 28° C. and was partitioned between water (25 mL) and EtOAc(40 mL×2). Organic layers were separated and was dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to get crude, then it waspurified by column chromatography (using 100-200 silica gel, columneluted at 80% ethyl acetate in hexane) to afford the(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(350 mg, 0.562 mmol, 19.88% yield) as an off white solid, LCMS (m/z):604.24 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

DIPEA (2.52 mL, 14.13 mmol) followed by triphosgene (0.503 g, 1.696mmol) were added to a solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1.0 g, 2.83 mmol) and(S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (0.828g, 3.67 mmol) in Tetrahydrofuran (THF) (50 mL) at 25° C., stirred for 16h at 70° C. Progress of the reaction was monitored by LCMS and TLC. Thereaction mixture was cooled to 28° C. and was partitioned between water(100 mL) and EtOAc (2×100 mL). Organic layer was separated and was driedover anhydrous Na₂SO₄, filtered and filtrate was evaporated to getcrude. The sample was loaded in dichloromethane and purified on silica(Si) 5 g using a 0-15% methanol-dichloromethane over 80 min. Theappropriate fractions were combined and evaporated in vacuum to give therequired product, 300 mg as a off-white solid, LCMS (m/z): 605.21[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(4-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

TEA (1.970 mL, 14.13 mmol) followed by triphosgene (0.839 g, 2.83 mmol)were added to a solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1 g, 2.83 mmol) in Tetrahydrofuran (THF) (30 mL) at RT and stirred for6 h at RT and(R)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (0.700g, 3.11 mmol) was added and heated at 80° C. for h. The reaction mixturewas cooled to 28° C. and was partitioned between water (25 mL) and EtOAc(40 mL×2). Organic layers were separated and was dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to get(9S)-3-chloro-N-(4-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.411 mmol, 14.52% yield) as a gum, LCMS (m/z): 605.26 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

DIPEA (329 mg, 2.54 mmol) followed by triphosgene (252 mg, 0.848 mmol)were added to a solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(300 mg, 0.848 mmol) in Tetrahydrofuran (THF) (50 mL) at 25° C., stirredfor 3 h and(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (380 mg,1.696 mmol) was added and stirred at 70° C. for 16 h. The reactionmixture was cooled to 28° C. and was partitioned between water (20 mL)and EtOAc (50 mL). Organic layer was separated and was dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to get crude. Thecrude mass was purified by column chromatography (100-200) mesh elutedwith 30% EtOAc in pet ether to obtained(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.414 mmol, 48.8% yield), as a pale yellow solid, LCMS (m/z):604.00[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

triphosgene (419 mg, 1.413 mmol) was added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.413 mmol), and TEA (0.985 mL, 7.07 mmol) in Tetrahydrofuran(THF) (20 mL) at 28° C. The reaction mixture was stirred for 30 min andwas added (S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine(634 mg, 2.83 mmol). The reaction mixture was stirred for 15.5 h at 70°C. The reaction mixture was cooled to room temperature and waspartitioned between water (30 mL) and EtOAc (100 mL). EtOAc layer wasseparated and was dried over anhydrous Na₂SO₄, filtered. The filtratewas evaporated to get crude. The sample was loaded in dichloromethaneand purified on silica (Si) 5 g using a 0-50% of Ethyl acetate/Pet etherover 80 mins. The appropriate fractions were combined and evaporated invacuum to give the required product 250 mg as an off-white solid. LCMS(m/z): 604.22 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To solid(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(600 mg, 1.696 mmol) in Tetrahydrofuran (THF) (20 mL) stirred undernitrogen at room temp was added TEA (1.182 mL, 8.48 mmol) and solidtriphosgene (302 mg, 1.018 mmol) stirred for 6 hrs and was added(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (571 mg,2.54 mmol). The reaction mixture was stirred at 80° C. for 16 hr. Thereaction mixture was monitored by TLC. The organic phase was evaporated,added water 50 mL and Extracted with Ethyl acetate and washed withsaturated brine 100 mL dried over Na₂SO₄ and evaporated in vacuum togive the crude products. The residue was purified via combiflash (100%ACN reverse phase column). Collected fractions and evaporated to getpure compound(9S)-3-chloro-N-(6-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(180 mg, 0.273 mmol, 16.12% yield), LCMS (m/z): 604.09 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

DIPEA (877 mg, 6.78 mmol) followed by triphosgene (671 mg, 2.261 mmol)were added to a solution of(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (1014 mg,4.52 mmol) in Tetrahydrofuran (THF) (25 mL) at 25° C., stirred for 1 hand(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(800 mg, 2.261 mmol) was added and heated at 70° C. for 18 hr. Thereaction mixture was cooled to 28° C. and was partitioned between water(20 mL) and EtOAc (50 mL). Organic layer was separated and was driedover anhydrous Na₂SO₄, filtered and filtrate was evaporated to get crude(TLC eluent: 5% methanol in DCM R_(f) 0.3; UV active). The crudecompound was purified by column chromatography (C-18: eluting with 80%ACN in 1% aq formic acid) to get 460 mg with LCMS: 73%. Further purifiedby flash column chromatography (silica-gel: 100-200 mesh) eluted with50% EtOAc in hexane to afford(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.490 mmol, 21.65% yield), as a brownish sticky, LCMS (m/z):604.0 (M+H)⁺.

Synthesis of(9S)-3-chloro-N-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To solid(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(600 mg, 1.696 mmol) in Tetrahydrofuran (THF) (20 mL) stirred undernitrogen at room temp was added TEA (1.182 mL, 8.48 mmol) and solidtriphosgene (302 mg, 1.018 mmol) stirred for 16 hrs and was added(S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (571 mg,2.54 mmol). The reaction mixture was stirred at 65° C. for 16 hr. Thereaction mixture was monitored by TLC. The organic phase was evaporatedadded water 50 mL and Extracted with Ethyl acetate and washed withsaturated brine 100 mL dried over Na₂SO₄ and evaporated in vacuum togive the crude products. The crude product purified by flashchromatography, collected fractions to get compound, was washed withdiethyl ether and pentane and filtered and washed pentane to get purecompound(9S)-3-chloro-N-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(216 mg, 0.319 mmol, 18.79% yield), LCMS (m/z): 604.11 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Procedure: triphosgene (377 mg, 1.272 mmol) was added to a stirredsolution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.272 mmol), and TEA (0.886 mL, 6.36 mmol) in Tetrahydrofuran(THF) (10.0 mL) at 28° C. The reaction mixture was stirred for 30 minand was added(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-amine (571 mg,2.54 mmol). The reaction mixture was stirred for 10 h at 70° C. Thereaction mixture was cooled to room temperature and was partitionedbetween water (5 mL) and EtOAc (15 mL). EtOAc layer was separated andwas dried over anhydrous Na₂SO₄, filtered. The filtrate was evaporatedto get crude. The crude was purified by chromatography (GRACE using C-18reserval column, Mobile phase A: 0.1% Formic Acid in water; B: MeOH,eluent 91% B in A). Combined fractions were concentrated basified withsaturated NaHCO₃. The aqueous layer was extracted with DCM, DCM layerwas dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated toafford(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300.0 mg, 0.489 mmol, 38.5% yield) as off-white solid, LCMS (m/z):604.13[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To solid(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(600 mg, 1.696 mmol) in Tetrahydrofuran (THF) (50 mL) stirred undernitrogen at room temp was added TEA (0.236 mL, 1.696 mmol) and solidtriphosgene (503 mg, 1.696 mmol) stirred for 30 min and was added(R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-amine (380 mg,1.696 mmol). The reaction mixture was stirred at 65° C. for 16 hr. Thereaction mixture was monitored by TLC. The organic phase was evaporatedadded water 50 mL and Extracted with Ethyl acetate and washed withsaturated brine 100 mL dried over Na₂SO₄ and evaporated in vacuum togive the crude products. The crude product was added to a silica gelcolumn and was eluted with DCM/MeOH. Collected fractions to get somepure compound again purified by via combiflash (100% ACN; 120 g reversephase column). Collected fractions to get pure compound(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(450 mg, 0.732 mmol, 43.2% yield), LCMS (m/z): 604.02 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (377 mg, 1.272 mmol) was added to a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(450 mg, 1.272 mmol) and TEA (1.064 mL, 7.63 mmol) in Tetrahydrofuran(THF) (30 mL) at 28° C. The reaction mixture was stirred for 2 h and wasadded (R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine(573 mg, 2.54 mmol). The reaction mixture was stirred for 16 hr at 65°C. TLC eluent: 100% Ethyl acetate R_(f): 0.3, UV active. The reactionmixture was cooled to room temp, solvent evaporated under reducedpressure completely and was partitioned between water (10 mL) and EtOAc(2×50 mL). Organic layer was separated, dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to give crude as brown solid. Crudewas diluted with DCM and absorbed with neutral alumina and eluted with35-40% EtOAc in pet ether fractions were collected and concentrated toget(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(220 mg, 0.244 mmol, 19.18% yield) as a brown solid, LCMS (m/z): 605.00[M+H]⁺.

Synthesis of(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

A solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(400 mg, 1.131 mmol), triphosgene (336 mg, 1.131 mmol) and triethylamine(0.788 mL, 5.65 mmol) in Tetrahydrofuran (THF) (20 mL) was stirred undernitrogen at room temp for 30 min. To this reaction mixture(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine (509mg, 2.261 mmol) was added. The reaction mixture was stirred at 70° C.for 16 h and progress of the reaction was monitored by TLC. The reactionmixture was cooled to room temperature, poured in to water (10 mL) andextracted with EtOAc (3×20 mL). The combined organic layer was washedwith water (20 mL), brine solution (20 mL), dried over Na₂SO₄, filteredand evaporated to get crude compound. TLC eluent: 100% EtOAc/Hexane,R_(f): 0.3, UV active. The crude compound was purified by columnchromatography using Neutral Alumina and eluted at 20% EtOAc in Petetherto afford pure(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.395 mmol, 34.9% yield) as off white solid, LCMS (m/z): 605.23[M+H

Synthesis of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a suspension of(9S)-2-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(50 g, 238 mmol), in Methanol (500 mL) at room temp was added TEA (100mL, 715 mmol). and PdCl₂(dppf)-CH₂Cl₂ adduct (9.74 g, 11.92 mmol). inautoclave was filled with CO (100 psi) gas. This was degassed and againfilled with CO (300-350 psi) gas. The reaction mixture was heated to130° C. for 16 hr. Progress of the reaction was monitored by TLC. TLCindicated starting material was consumed. Cooled the reaction mass toroom temperature, filtered through celite, washed the celite bed withMethanol (500 mL). The filtrate was concentrated to get black stickycompound. To this added Methanol (200 mL) and stirred for 15 minutes andfiltered the solid and dried to afford (9 S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(30 g, 108 mmol, 45.1% yield)) (N36489-30-A2) as Off-white solid, LCMS(m/z): 234.07 (M+H)⁺.

Synthesis of(9S)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(3 g, 12.86 mmol) in Tetrahydrofuran (THF) (20 mL), Water (20 mL). Thereaction mixture was cooled to 0° C. and added a solution of LiOH (0.462g, 19.29 mmol) in Water (10 mL). The reaction was stirred at 28° C. for2 h and progress of the reaction was monitored by TLC. The solvent wasevaporated under reduced pressure, diluted with water and acidified with1N HCl (pH; 4-5) at 0° C. The solid was filtered, washed with water anddried under vacuum to afford(9S)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (2.0 g, 8.54 mmol, 66.4% yield) as an off white solid, LCMS (m/z):220.00 [M+H]⁺.

Synthesis of(9S)-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b]diazocine-2-carboxamide

To a stirred solution of(9S)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (2.0 g, 9.12 mmol), HATU (5.20 g, 13.68 mmol) inN,N-Dimethylformamide (DMF) (20 mL) was added DIPEA (6.37 mL, 36.5mmol). (R)-1,1,1-trifluoropropan-2-amine (1.032 g, 9.12 mmol) was addedto the reaction mixture at 0° C. The reaction mixture was stirred for 16hr at 28° C. and progress of the reaction was monitored by TLC. Thereaction mixture was poured in to water (40 mL) and extracted with EtOAc(3×30 mL). The combined organic layer was washed with brine solutiondried over anhydrous Na₂SO₄, filtered and evaporated to get crudecompound. The crude compound was purified by column chromatography usingneutral alumina and eluent at 30% EtOAc in Pet ether to afford(9S)-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxamide(1.8 g, 5.59 mmol, 61.3% yield) as an off white solid, LCMS (m/z):314.85 [M+H]⁺.

Synthesis of(9S)-N10-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxamide(300 mg, 0.954 mmol), triphosgene (283 mg, 0.954 mmol) and triethylamine(0.665 mL, 4.77 mmol) in Tetrahydrofuran (THF) (20 mL) under nitrogen at28° C. and the reaction mixture was stirred at rt for 30 min.(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (322 mg,1.432 mmol) was added to the reaction mixture. The reaction mixture wasstirred for 16 hr at 70° C. and progress of the reaction was monitoredby TLC. The reaction mixture was cooled to rt, partitioned between water(30 mL) and EtOAc (3×30 mL).

Organic layer was separated and was dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to give crude. The crude waspurified by GRACE using C-18 reserval column, Mobile phase A: 0.1%Formic Acid in water; B: MeOH, the product was eluted at 65% of MeOH and0.1% Formic Acid in water. The solvent was evaporated and basified withsaturated NaHCO₃. The precipitated solid was filtered, and was dried toafford(9S)-N10-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(200 mg, 0.349 mmol, 36.6% yield) as an off-white solid, LCMS (m/z):566.35 [M+H]⁺.

Synthesis of(9S)-10-((5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a stirred solution of (9S)-methyl10-((5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(400 mg, 0.827 mmol) in Tetrahydrofuran (THF) (10 mL), Water (10 mL) wasadded a solution of LiOH (29.7 mg, 1.241 mmol) in water (1 mL) at 0° C.The reaction mixture was stirred for 2 h at 28° C. and progress of thereaction was monitored by TLC. The reaction solvent was evaporated underreduced pressure, diluted with water and acidified with 1N HCl (pH; 2-3)at 0° C. and extracted with DCM (3×40 mL). The combined organic layerwas washed with water, brine solution dried over Na₂SO₄ Filtered andevaporated to get crude compound. The crude was purified by GRACE usingC-18 reserval column, Mobile phase A: 0.1% Formic Acid in water; B:MeOH, the product was eluted at 50% of MeOH and 0.1% Formic Acid inwater. The solvent was evaporated and basified with saturated NaHCO₃.The precipitated solid was filtered and was dried to afford(9S)-10-((5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (350 mg, 0.802 mmol, 97% yield) as an off-white solid, LCMS (m/z):430.46 [M+H]⁺.

Synthesis of(9S)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (500 mg, 1.065 mmol) and (R)-1,1,1-trifluoropropan-2-amine (241 mg,2.130 mmol) in Tetrahydrofuran (THF) (10 mL) stirred under nitrogen at28° C. was added HATU (486 mg, 1.278 mmol) and DIPEA (0.372 mL, 2.130mmol) and the reaction mixture was stirred at 28° C. for 16 hr. Reactionmixture was quenched with ice water and extracted with 3×50 ml of ethylacetate, combined organic layers were washed with 100 ml of brinesolution and dried over Na₂SO₄ and concentrated under reduced pressureto afford crude compound. The crude compound was purified by columnchromatography (100-200 silica gel) using gradient mixture of 80% EtOAcin Petether as eluent, to afford the(9S)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(200 mg, 0.341 mmol, 32.0% yield) as an off white solid, LCMS (m/z):565.15 [M+H]⁺.

Synthesis of(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1.0 g, 2.068 mmol) in Tetrahydrofuran (THF) (35 mL) and Water (35.0 mL)was added LiOH (0.074 g, 3.10 mmol). The reaction mixture was stirred atRT for 1 hr. Reaction mixture was concentrated under reduced pressure toafford compound(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (900 mg, 1.876 mmol, 91% yield) as Off white solid, LCMS (m/z):470.16 (M+H)⁺.

Synthesis of (9S)-methyl10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a solution of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1.0 g, 4.29 mmol), triphosgene (0.763 g, 2.57 mmol) in Tetrahydrofuran(THF) (30 mL) stirred under nitrogen at 0° C. and added DIPEA (3.74 mL,21.43 mmol). Then the reaction mixture was stirred at 28° C. for 30 minand added (S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine(1.442 g, 6.43 mmol), then the reaction mixture was stirred at 80° C.for 15.5 hr. The reaction was monitored by LCMS and TLC. The reactionmixture was poured in to the cold water (50 mL) and extracted with ethylacetate (2×100 mL). The organic layer was dried over anhydrous Na₂SO₄and concentrated under vacuum to give crude product, LCMS (m/z): 484.14(M+H)⁺.

Synthesis of (9S)-methyl10-((4-bromopyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a solution of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1.0 g, 4.29 mmol), TEA (3.59 mL, 25.7 mmol) in Tetrahydrofuran (THF)(30 mL) stirred under nitrogen at 0° C. and added triphosgene (1.272 g,4.29 mmol). Then the reaction mixture was stirred at 30° C. for 30 minand added 4-bromopyridin-2-amine (2.225 g, 12.86 mmol), then thereaction mixture was stirred at 80° C. for 15.5 hr. The reaction wasmonitored by LCMS and TLC. The reaction mixture was poured in to thecold water (20 mL) and extracted with ethyl acetate (2×50 mL). Theorganic layer was dried over anhydrous Na₂SO₄ and concentrated undervacuum to give crude product, LCMS (m/z): 431.9 (M+H)⁺.

Synthesis of (9S)-methyl10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate;and(9S)-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a suspension of NaH (8.57 g, 214 mmol) in Tetrahydrofuran (THF) (100mL), stirred under nitrogen at room temperature, was added solid(9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(10 g, 42.9 mmol), after 15 min at room temperature was added3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (15.45 g,64.3 mmol). The resulting reaction mixture was stirred at 80° C. for 4hr. Progress of the reaction was monitored by TLC. TLC indicatedstarting material was consumed. The reaction mass was cooled to rt,diluted with Ethyl acetate (100 mL), quenched in ice-cold water (250mL). Separated the organic layer and was dried over Na₂SO₄, filtered andconcentrated to get (9S)-methyl10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(10 g, 11.35 mmol, 26.5% yield) as brown solid compound. LCMS (m/z):354.16 (M+H)⁺. The aqueous layer was acidified with 2N HCl to pH 5 to 6.The aqueous layer was concentrated under vacuum to get the off whitesolid compound. This was dissolved in 10% Methanol in DCM (1.0 L) andfiltered the inorganic. The filtrate was concentrated and dried toafford(9S)-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (3 g, 5.46 mmol, 12.75% yield) as light brown solid. LCMS (m/z):340.0 (M+H)⁺.

Synthesis of (9S)-methyl3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a suspension of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(2.2 g, 9.43 mmol), in Chloroform (25 mL) stirred under nitrogen at 0°C., was added 1-chloropyrrolidine-2,5-dione (1.637 g, 12.26 mmol) in lotwise over 2 min at 0° C. After 30 min at 0° C. reaction mixture stirredat room temperature for 3 hr. Progress of the reaction was monitored byTLC. TLC indicated two non polar spots and small amount of un-reactedSM. Reaction mass was diluted with 50 ml of water, extracted with (2×100ml) of DCM. Combined organic layers were dried over Na₂SO₄, filtered andconcentrated to get crude. Crude material was purified by combiflashusing silica gel column (40 g, 50% EtOAc in Hexane). Fractionscontaining pure compound were combined and concentrated to afford thedesired compound (9S)-methyl3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1 g, 3.29 mmol, 34.9% yield) as pale brown color viscous liquid, LCMS(m/z): 267.96 (M+H)⁺.

Synthesis of (9S)-methyl10-(isoxazol-3-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a solution of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(600 mg, 2.57 mmol), triphosgene (458 mg, 1.543 mmol) in Tetrahydrofuran(THF) (10 mL) stirred under nitrogen at 0° C. and added DIPEA (2.246 mL,12.86 mmol). Then the reaction mixture was stirred at 30° C. for 30 minand added isoxazol-3-amine (324 mg, 3.86 mmol), then the reactionmixture was stirred at 80° C. for 15.5 hr. The reaction was monitored byLCMS and TLC. The reaction mixture was poured in to the cold water (50mL) and extracted with ethyl acetate (2×50 mL). The organic layer wasdried over anhydrous Na₂SO₄ and concentrated under vacuum to give crudeproduct, LCMS (m/z): 344.17 (M+H)⁺.

Synthesis of(9S)-10-(isoxazol-3-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl10-(isoxazol-3-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(0.5 g, 1.456 mmol) in Tetrahydrofuran (THF) (25 mL) and Water (25.00mL) was added LiOH (0.052 g, 2.184 mmol). The reaction mixture wasstirred at RT for 1 hr. Reaction mixture was concentrated under reducedpressure to remove all the THF and then acidified with 2N.HCl up topH=4. then the precipitated solid filtered and dried to afford compound(9S)-10-(isoxazol-3-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (450 mg, 1.346 mmol, 92% yield) as an off white solid, LCMS (m/z):330.05 (M+H)⁺.

Synthesis of (9S)-methyl10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a solution of (9S)-methyl7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(2 g, 8.57 mmol) in stirred under nitrogen at 0° C. triphosgene (2.54 g,8.57 mmol) and TEA (7.17 mL, 51.4 mmol) was added. Then the reactionmixture was stirred at 30° C. for 30 min and added6-methyl-1H-pyrazolo[3,4-b]pyridin-3-amine (507 mg, 3.42 mmol), then thereaction mixture was stirred at 90° C. for 16 hr. The reaction wasmonitored by LCMS and TLC. The reaction mixture was poured in to thecold water (30 mL) and extracted with ethyl acetate (2×50 mL). Theorganic layer was dried over anhydrous Na₂SO₄ and concentrated undervacuum to obtained crude compound. The crude compound was purified bycolumn chromatography (100-200 silica gel) using gradient mixture of 10%methanol in DCM as eluent to afford the compound (9S)-methyl10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1.5 g, 2.338 mmol, 27.3% yield) as pale brown solid, LCMS (m/z): 408.00(M+H)⁺.

Synthesis of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1.5 g, 3.68 mmol) in Tetrahydrofuran (THF) (10 mL) and Water (10.00 mL)was added LiOH (0.132 g, 5.52 mmol). The reaction mixture was stirred atRT for 2 hr. Reaction mixture was concentrated under reduced pressure toafford compound as Li salt(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (1.2 g, 3.03 mmol, 82% yield) as Pale brown solid, LCMS (m/z):394.1 (M+H)⁺.

Synthesis of(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a stirred solution of NaH (0.448 g, 18.68 mmol) in Tetrahydrofuran(THF) (80 mL) at room temperature, was added (9S)-methyl3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(1 g, 3.74 mmol) in portion wise during 1 min, After 15 min at roomtemperature was added3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (1.077 g,4.48 mmol). The resulting reaction mixture was stirred at roomtemperature for 1 hr. And then at 70° C. for 3 hr. Progress of thereaction was monitored by TLC. TLC indicated formation of a non polarspot, a polar spot and complete consumption of SM. Reaction mass wasdiluted with 100 ml of ice cold water, aqueous layer was washed with(100 ml) of EtOAc, pH of aqueous layer adjusted to 4 with 1N HCl,concentrated aqueous layer under reduced pressure to get crude,resulting crude solid was extracted with 10% MeOH in DCM andconcentrated the organic layer to get(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (500 mg, 0.744 mmol, 19.91% yield), LCMS (m/z): 374.20 (M+H)⁺.

Synthesis of(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a stirred solution of NaH (224 mg, 5.60 mmol) in Tetrahydrofuran(THF) (30 mL) at room temperature, was added (9S)-methyl3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(500 mg, 1.868 mmol) in portion wise during 1 min, After 15 min at roomtemperature was added3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (538 mg,2.241 mmol). The resulting reaction mixture was stirred at roomtemperature for 1 hr. And then at 70° C. for 3 hr. Progress of thereaction was monitored by TLC. TLC indicated formation of a non polarspot, a polar spot and complete consumption of SM. Reaction mass wasdiluted with 100 ml of ice cold water, aqueous layer was washed with(100 ml) of EtOAc, pH of aqueous layer adjusted to 4 with 1N HCl,concentrated aqueous layer under reduced pressure to get crude,resulting crude solid was extracted with 10% MeOH in DCM andconcentrated the organic layer to get(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (300 mg, 0.525 mmol, 28.1% yield) N35384-85-A1 as Brown color, LCMS(m/z): 374.08 (M+H)⁺.

Synthesis of(9S)-3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(2.0 g, 7.47 mmol) in Tetrahydrofuran (THF) (35 mL) and Water (35.0 mL),LiOH (0.268 g, 11.21 mmol) was added at RT and stirred for 16 hrs. (TLCeluent: 10% MeOH in ethyl acetate R_(f): 0.2; UV active). Reactionmixture was diluted with water (100 mL), extracted with ethyl acetate(2×100 mL) to remove all the impurities. The aqueous layer acidifiedwith aq.HCl (5 mL) and concentrated under reduced pressure to afford(9S)-3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (1.02 g, 3.32 mmol, 44.4% yield) as an light brown colored solid.LCMS (m/z): 253.91 [M+H]⁺.

Synthesis of(9S)-3-chloro-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxamide

To a solution of(9S)-3-chloro-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (1.02 g, 4.02 mmol) in N,N-Dimethylformamide (DMF) (5 mL) undernitrogen at room temp, HATU (1.835 g, 4.82 mmol), DIPEA (1.404 mL, 8.04mmol) and (R)-1,1,1-trifluoropropan-2-amine (0.682 g, 6.03 mmol) wasadded and stirred at RT for 16 h. Reaction mixture was diluted with icewater and extracted with 2×100 mL of ethyl acetate. The combined organiclayers were washed with brine, dried over Na₂SO₄ and concentrated underreduced pressure to afford crude compound. The crude product waspurified by flash column chromatography (100-200 silica gel eluted with5% of CH₂Cl₂/MeOH) to afford(9S)-3-chloro-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxamide(800 mg, 1.991 mmol, 49.5% yield) as an off white solid, LCMS (m/z):348.96 [M+H]⁺.

Synthesis of(9S)-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(10 g, 28.3 mmol), and in Tetrahydrofuran (THF) (100 mL), Water (100 mL)at room temp was added lithium hydroxide (1.355 g, 56.6 mmol). Thereaction mixture was stirred at rt for 3 hr. Progress of the reactionwas monitored by TLC. TLC indicated starting material was consumed. Thereaction mass was diluted with Ethyl acetate (100 mL), separated theorganic layer. The aqueous layer was acidified with 1N HCl to PH-5-6.The aqueous layer was concentrated to get light brown solid. To thisadded 10% Methanol in DCM (500 mL) and stirred for 15 minutes andfiltered the inorganic. The filtrate was concentrated to afford(9S)-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (6 g, 17.47 mmol, 61.7% yield) as light brown solid. LCMS (m/z):340.15 (M+H)⁺.

Synthesis of(9S)-10-((4-bromopyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a solution of (9S)-methyl10-((4-bromopyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(0.800 g, 1.851 mmol) in Tetrahydrofuran (THF) (35 mL) and Water (35.0mL) was added LiOH (0.066 g, 2.78 mmol). The reaction mixture wasstirred at RT for 1 hr. Reaction mixture was concentrated under reducedpressure to afford Li salt of the product. Then the salt was dilutedwith water (20 mL) washed thoroughly with Ethyl acetate (2×50 mL) thenthe aqueous layer acidified with aq. HCl to afford(9S)-10-((4-bromopyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (600 mg, 1.357 mmol, 73.3% yield) as an Off white solid, LCMS(m/z): 419.90 [M+H]⁺.

Synthesis of(9S)-N10-(4-bromopyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-10-((4-bromopyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (480 mg, 1.148 mmol) and (R)-1,1,1-trifluoropropan-2-amine (195 mg,1.721 mmol) in N,N-Dimethylformamide (DMF) (15 mL) stirred undernitrogen at 28° C. was added HATU (524 mg, 1.377 mmol) and DIPEA (0.401mL, 2.295 mmol) and the reaction mixture was stirred at 28° C. for 16hr. Reaction mixture was quenched with ice water and extracted with 3×20ml of ethyl acetate, combined organic layers were washed with 20 ml ofbrine solution and dried over Na₂SO₄ and concentrated under reducedpressure to afford crude compound. Crude compound was purified by columnchromatography using 100-200 silica gel and with an eluent 80-100% ofethyl acetate/Hexane. Collected fractions were distilled off to afford(9S)-N10-(4-bromopyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(500 mg, 0.960 mmol, 84% yield) as an off white solid, LCMS (m/z): 565.9[M+H]⁺.

Synthesis of(9S)-N2-(2,2-difluoropropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (500 mg, 1.065 mmol) in Tetrahydrofuran (THF) (10 mL) stirred undernitrogen at 20° C. was added HATU (486 mg, 1.278 mmol) and DIPEA (0.372mL, 2.130 mmol) and the reaction mixture was stirred at 28° C. for 16hr. Reaction mixture was diluted with ice water and extracted with 3×50ml of ethyl acetate, combined organic layers were washed with 100 ml ofbrine solution and dried over Na₂SO₄ and concentrated under reducedpressure to afford crude compound. Crude compound was purified by columnchromatography using 100-200 mesh silicagel and was eluted with 80% toPure ethyl acetate, LCMS (m/z): 547.35 [M+H]⁺.

Synthesis of:(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(Peak-1); and(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(Peak-2)

To a stirred solution of(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (2.5 g, 5.32 mmol) in ACN (10 mL) was added HATU (4.05 g, 10.65mmol) and DMAP (1.301 g, 10.65 mmol). Reaction mixture was stirred at RTfor 15 min, then added 2,2-difluorocyclopropanamine, Hydrochloride(1.380 g, 10.65 mmol) in sealed tube, reaction mixture was stirred at90° C. for 16 hr. Progress of the reaction was monitored by TLC.Reaction mixture was cooled to RT, Water (50 mL) and extracted withethyl acetate (2×50 mL), separated organic layer, dried over Na₂SO₄,concentrated under reduced pressure to obtain crude. Obtained crude waspurified by column using silica (100-200 mesh/1-80% EtOH in Pet-ether asa eluent), collected fractions were concentrated pressure to get purecompound 700 mg. The obtained pure compound was purified by SFC.

After SFC purification to get(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(Peak-1) (230 mg, 0.414 mmol, 7.77% yield) and(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(Peak-2) (250 mg, 0.454 mmol, 8.53% yield).

Column/dimensions: Chiralpak IC (250×30) mm, 5μ % CO2: 50.0%

% Co solvent: 50.0% (100% Methanol)Total Flow: 100.0 g/min

Back Pressure: 100.0 bar UV: 210 nm

Stack time: 6.5 min

Load/Inj: 9.5 mg Solubility: Methanol

Total No of injections: 70Instrument details: Make/Model: Thar SFC-200-NEW-2LCMS (m/z): 545.02 [M+H]t (peak 1)LCMS (m/z): 545.89 [M+H]t (peak 2)

Synthesis of(9S)-N10-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (400 mg, 0.827 mmol) in N,N-Dimethylformamide (DMF) (8 mL) wereadded DIPEA (0.722 mL, 4.14 mmol), HATU (472 mg, 1.241 mmol) and(R)-1,1,1-trifluoropropan-2-amine (140 mg, 1.241 mmol) at 0° C. Thereaction mixture was stirred at 27° C. for 3 hr. The reaction mixturewas diluted with cold water (100 mL) and stirred for 15 min. Theprecipitated solid was filtered through Buchner Funnel, washed withwater and dried under reduced pressure to get crude compound. The crudecompound was purified by column chromatography (neutral alumina, eluent:65-70% ethyl acetate in hexane). Collected fractions were concentratedunder reduce pressure to afford desired product(9S)-N10-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(350 mg, 0.580 mmol, 70.1% yield) as a brown solid, LCMS (m/z): 579.22[M+H]⁺.

Synthesis of(9S)-10-((5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a stirred solution of (9S)-methyl10-((5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(450 mg, 0.904 mmol) in Tetrahydrofuran (THF) (20 mL) & Water (3.0 mL)was added LiOH.H₂O (21.66 mg, 0.904 mmol) at 0° C. The reaction mixturewas stirred at 27° C. for 6 hr. The reaction mixture solvent wasevaporated under reduced pressure completely and acidified with citricacid solution at 0° C. The aqueous layer was extracted withdichloromethane (2×50 mL). Organic layer was separated, dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude asbrown solid. The crude compound was triturated with diethyl ether andpentane, LCMS (m/z): 484.28 [M+H]⁺.

Synthesis of (9S)-methyl10-((5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a stirred solution of (9S)-methyl4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(600 mg, 2.426 mmol) in Tetrahydrofuran (THF) (45 mL) were added TEA(2.029 mL, 14.56 mmol) and triphosgene (720 mg, 2.426 mmol) undernitrogen at 28° C. The reaction mixture was stirred at rt for 30 min,then (R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine(1088 mg, 4.85 mmol) was added to the reaction mixture. The reactionmixture was stirred 16 hr at 65° C. The reaction mixture was cooled toroom temp, solvent evaporated under reduced pressure completely and waspartitioned between water (30 mL) and EtOAc (2×40 mL). Organic layer wasseparated, dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to give crude as brown solid. TLC eluent: 100% EtOAc/Hexane,R_(f): 0.3, UV active. The crude compound was purified by Grace usingC-18 reserval column, Mobile phase A: 0.1% Formic Acid in water; B: ACN,the product was eluted at 70-75% ACN/0.1% Formic Acid in water. Thesolvent was evaporated and was basified with saturated NaHCO₃. Theaqueous layer was extracted with DCM. DCM layer was dried over anhydrousNa₂SO₄, filtered and evaporated to afford pure (9S)-methyl10-((5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(500 mg, 0.936 mmol, 38.6% yield) as a off white solid, LCMS (m/z):498.48 [M+H]⁺.

Synthesis of (9S)-methyl4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a stirred solution of(9S)-2-chloro-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(6 g, 26.8 mmol) in Methanol (100 mL) was degassed for 30 min, thentriethylamine (18.69 mL, 134 mmol) and PdCl₂(dppf)-CH₂Cl₂ adduct (1.095g, 1.341 mmol) were added and filled with 300 psi CO gas. The reactionmixture was stirred at 115° C. for 16 h in steel bomb. The reaction wasmonitored by TLC. The reaction mixture was concentrated under reducedpressure to afford crude compound. The crude product was dissolved inDCM (50 mL), and was washed with water (10 mL). The DCM layer was driedover anhydrous Na₂SO₄, filtered, and filtrate was evaporated to afford(9 S)-methyl4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(7 g, 23.76 mmol, 89% yield) as a brown solid, LCMS (m/z): 248.11[M+H]⁺.

Synthesis of(9S)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (450 mg, 0.931 mmol) in N,N-Dimethylformamide (DMF) (8 mL) wereadded DIPEA (0.813 mL, 4.65 mmol), HATU (531 mg, 1.396 mmol) and(R)-1,1,1-trifluoropropan-2-amine (158 mg, 1.396 mmol) at 0° C. Thereaction mixture was stirred at 27° C. for 3 hr. The reaction mixturewas diluted with cold water (100 mL) and stirred for 15 min. Theprecipitated solid was filtered through Buchner Funnel, washed withwater and dried under reduced pressure to get crude compound. Thecompound was triturated with ether and to afford(9S)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(330 mg, 0.481 mmol, 51.7% yield) as an off white solid, LCMS (m/z):579.32 [M+H]⁺.

Synthesis of(9S)-10-(((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid

To a stirred solution of (9S)-methyl10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(550 mg, 1.105 mmol) in Tetrahydrofuran (THF) (20.0 mL) & Water (5.00mL) was added LiOH.H₂O (39.7 mg, 1.658 mmol) at 0° C. The reactionmixture was stirred at 27° C. for 6 hr. The reaction mixture solvent wasevaporated under reduced pressure completely and acidified with citricacid solution at 0° C. The aqueous layer was extracted withdichloromethane (2×50 mL). Organic layer was separated, dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude.The crude compound was triturated with diethyl ether and pentane to get(9S)-10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (450 mg, 0.892 mmol, 81% yield) as a off white solid, LCMS (m/z):484.30 [M+H]⁺.

Synthesis of (9S)-methyl10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate

To a stirred solution of (9S)-methyl4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(800 mg, 3.24 mmol) in Tetrahydrofuran (THF) (55 mL) were added TEA(2.71 mL, 19.41 mmol) and triphosgene (960 mg, 3.24 mmol) under nitrogenat 28° C. The reaction mixture was stirred at rt for 30 min, then(S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (2176 mg,9.71 mmol) was added to the reaction mixture. The reaction mixture wasstirred 16 hr at 65° C. The reaction mixture was cooled to room temp,solvent evaporated under reduced pressure completely and was partitionedbetween water (30 mL) and EtOAc (2×40 mL). Organic layer was separated,dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated togive crude as brown solid. TLC eluent: 100% EtOAc/Hexane, R_(f):0.2, UVactive. The crude compound was purified by Grace using C-18 reservalcolumn, Mobile phase A: 0.1% Formic Acid in water; B: ACN, the productwas eluted at 65-70% ACN/0.1% Formic Acid in water. The solvent wasevaporated and was basified with saturated NaHCO₃. The aqueous layer wasextracted with DCM. DCM layer was dried over anhydrous Na₂SO₄, filteredand evaporated to afford pure (9S)-methyl10-((4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)carbamoyl)-4-methyl-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylate(650 mg, 1.125 mmol, 34.8% yield) as a off white solid, LCMS (m/z):498.48 [M+H]⁺.

Synthesis of(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine

To a stirred suspension of NaH (11.67 g, 292 mmol) inN-Methyl-2-pyrrolidone (NMP) (100 mL) under nitrogen at 0° C. was addeda solution of (R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (25.7 g, 194mmol) in N-Methyl-2-pyrrolidone (NMP) (100 mL) dropwise during 10 min at0° C. After 10 min added a solution of 6-chloropyridin-2-amine (25 g,194 mmol) in N-Methyl-2-pyrrolidone (NMP) (100 mL) dropwise during 10min at 0° C. The reaction mixture was heated at 100° C. for 36 hr. TLCindicates small amount starting material along with product.

Reaction mixture was poured into ice cold water (600 mL), aqueous layerwas extracted with EtOAc (2×500 mL). The organic layer was washed withwater (3×300 mL) to remove excess NMP. The organic layer was dried overNa₂SO₄, filtered and concentrated under reduced pressure to obtain crudeproduct. Crude product was purified by column chromatography using100-200 silica gel as a eluent (12-15% EtOAc in petether) to obtain(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (10 g,44.6 mmol, 22.93% yield) as a yellow thick liquid.

Synthesis of(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine

To a stirred suspension of NaH (62.2 g, 1556 mmol) inN-Methyl-2-pyrrolidone (NMP) (800 mL), under nitrogen at 0° C., wasadded a solution of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (206 g,1556 mmol) in N-Methyl-2-pyrrolidone (NMP) (300 mL) dropwise during 2 h.After stirring for another 10 min added a solution of6-chloropyridin-2-amine (200 g, 1556 mmol) in N-Methyl-2-pyrrolidone(NMP) (300 mL) dropwise during 30 min at 0° C. The reaction mixture wasstirred at 120° C. for 48 hr. TLC indicated that starting material was.Reaction mixture was poured into ice cold water (2000 mL), aqueous layerwas extracted with EtOAc (3×1000 mL). The combined organic layer waswashed with water (3×1000 mL) to remove excess NMP. The organic layerwas dried over Na₂SO₄, filtered and concentrated under reduced pressureto obtain crude product. Crude product was purified by columnchromatography using 100-200 silica gel (eluent 12-15% EtOAc in petether) to obtain the desired pure product(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (75 g,325 mmol, 20.92% yield) as a yellow viscous liquid. LCMS (m/z): 225[M+H]⁺.

Synthesis of(R)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine

To a suspension of (R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (3.000g, 22.70 mmol), 4-chloropyridin-2-amine (1.459 g, 11.35 mmol) and sodium(0.522 g, 22.70 mmol) in a sealed tube. The reaction mixture was stirredat 140° C. for 16 h. Next, the reaction mixture was cooled to roomtemperature, dissolved in MeOH and poured in to ice water and extractedwith EtOAc. The organic phase was washed with brine solution and driedover sodium sulfate, filtered and evaporated to get crude compound. Thecrude compound was purified by column chromatography using silica geland eluted with 2-3% MeOH/DCM to get pure compound (1.1 g, 21%), LCMS(m/z) 225.2 [M+H]⁺.

Synthesis of(S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine

To a suspension of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (3.000g, 22.70 mmol), 4-chloropyridin-2-amine (1.459 g, 11.35 mmol) and sodium(0.522 g, 22.70 mmol) in a sealed tube. The reaction mixture was stirredat 140° C. for 16 h before being cooled to room temperature, dissolvedin MeOH and poured in to ice water and extracted with EtOAc. The organicphase was washed with brine solution and dried over sodium sulfate,filtered and evaporated. The crude material was purified by silica gelcolumn chromatography eluting with 2-3% MeOH/DCM to give the desiredproduct (1.2 g, 22%), LCMS (m/z) 225.2 [M+H]⁺.

Synthesis of(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine

To a solution of 6-chloropyrazin-2-amine (5 g, 38.6 mmol), sodiumhydride (2.316 g, 57.9 mmol) and(R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (5.61 g, 42.5 mmol) inTetrahydrofuran (THF) (50 mL) stirred under nitrogen at 0° C. was addedreaction mixture was stirred at 80° C. for 16 h. Reaction mixture wasquenched with ice cold water and extracted into ethyl acetate. Organiclayer dried over Na₂SO₄. Solvent evaporated under reduced pressure toafford the crude product. The crude product was added to a silica gelcolumn and was eluted with DCM/MeOH. Fractions with product werecombined and evaporated under reduced pressure to give the requiredproduct (2.8 g, 11.9 mmol, 31%), LCMS (m/z) 225.9 [M+H]⁺.

Synthesis of(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine

6-chloropyrazin-2-amine (0.980 g, 7.57 mmol),(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (2 g, 15.13 mmol) andsodium (0.348 g, 15.13 mmol) were taken in a seal tube and heated at130° C. for 16 hr and then the reaction mixture was quenched withmethanol and ice cold water (100 mL) and extracted with ethyl acetate(5×50 mL). The combined organic layers were washed with water, saturatedbrine solution, dried over anhydrous sodium sulfate, filtered andconcentrated to give the product (1 g, 4.26 mmol, 28.2% yield), LCMS(m/z) 265.1 [M+H]⁺.

Synthesis of(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine

To suspension of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (10.20 g,77 mmol), and NaH (4.63 g, 116 mmol) in tetrahydrofuran (THF) (50 mL)stirred under nitrogen at room temperature was added2-chloropyrimidin-4-amine (5 g, 38.6 mmol) portion wise over 15 min. Thereaction mixture was stirred at 70° C. for 16 hr. Next, the reactionmixture was quenched with solution of aq. NaHCO₃ and then extracted withEtOAc, dried Na₂SO₄ and evaporated. The crude product was added to asilica gel column and was eluted with 50% Hex/EtOAc. Collected fractionswere evaporated to give the desired product (3 g, 11.84 mmol, 30.7%yield) as off white solid, LCMS (m/z) 226.2 [M+H]⁺.

Synthesis of(R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine

To a solution of sodium hydride (0.817 g, 34.1 mmol) in Tetrahydrofuran(THF) (30 mL) at room temperature was added a solution of(R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (3 g, 22.70 mmol) in THF (5mL) over 1 min and stirred at room temperature for 15 min then add2-chloropyrimidin-4-amine (2.059 g, 15.89 mmol) portion wise at roomtemperature. The reaction mixture was stirred at 65° C. for 16 h. Thereaction mixture was poured in to water and extracted with EtOAc (3×100mL). Then the combined organic layer was washed with water, brinesolution, dried over sodium sulfate and evaporated to get 4.0 g of crudecompound. The crude compound was purified by column chromatography using100-200 silica gel mesh and eluted with 2-3% MeOH/DCM to get purecompound (2.5 g, 10.42 mmol, 46%), LCMS (m/z) 226.2 [M+H]⁺.

Synthesis of(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-amine

To a suspension of 2-chloropyridin-4-amine (1.459 g, 11.35 mmol),(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (3.0 g, 22.70 mmol) wasadded sodium (0.522 g, 22.70 mmol). The reaction mixture was stirred at140° C. for 16 hr and progress of the reaction was monitored by

The reaction mixture was dissolved in MeOH, poured in to ice water andextracted with EtOAc (3×100 mL). Then the combined organic layer waswashed with water, brine solution, dried over sodium sulfate andevaporated to get 4.0 g of crude compound. The crude compound waspurified by column chromatography using 100-200 silica gel mesh andeluted with 2-3% MeOH/DCM to get(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-amine (2.5 g,10.73 mmol, 47.3% yield), LCMS (m/z) 225.3 [M+H]⁺.

Synthesis of(R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-amine

To a solution of 2-chloropyridin-4-amine (4 g, 31.1 mmol),(R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (2.056 g, 15.56 mmol) andsodium (0.715 g, 31.1 mmol) in sealed tube at room temperature. Thereaction mixture was stirred at 140° C. for 48 hr. The reaction mixturewas cooled to room temp and quenched with MeOH followed by water. Thenreaction mass was extracted with the EtOAc. Then organic layer washedwith water followed by brine solution and dried out with sodium sulfateand filtered and distill out completely. The crude product was added toa silica gel column and was eluted with Hex/EtOAc (1:1) collectedfractions were evaporated to give the desired product (2.250 g, 9.93mmol, 31.9% yield), LCMS (m/z) 225.0 [M+H]⁺.

Synthesis of phenyl (1-methyl-1H-pyrazol-4-yl)carbamate

Phenyl carbonochloridate (2.90 g, 18.53 mmol) was added to a stirredsolution of pyridine (3.12 mL, 38.6 mmol) in Dichloromethane (DCM) (50mL) at 0° C. and stirred for 15 min and followed by addition of1-methyl-1H-pyrazol-4-amine (1.5 g, 15.45 mmol) at same temperature. Thereaction mixture was stirred at room temperature for 4 h. Afterconsumption of starting material (monitored by TLC), ice cold water wasadded, separated organic layer was washed with water and brine. Theorganic layer was filtered through sodium sulfate and concentrated toget crude compound. The crude compound was purified by columnchromatography by using 60-120(silica gel) and eluted in 50% ethylacetate in hexane to afford the desired product (1.6 g, 6.41 mmol, 42%yield) as light brown solid, LCMS (m/z) 218.1 (M+H)⁺.

Synthesis of phenyl pyridin-3-ylcarbamate

To a solution of phenyl carbonochloridate (2.163 g, 13.81 mmol), andpyridine (1.375 mL, 17.00 mmol) in Dichloromethane (DCM) (30 mL) stirredunder nitrogen at room temp was added pyridin-3-amine (1.0 g, 10.63mmol). The reaction mixture was stirred at RT for 30 min. The reactionmixture was quenched with saturated sodium bicarbonate solution.Separated organic layer and the aqueous layer extracted with DCM (50ml). Combined DCM layer washed with water and dried out with sodiumsulfate, filtered and concentrated under high vacuum to get crudeproduct. The Crude product was added to a silica gel column and waseluted with 20% EtOAc/Hexane. Collected fractions were evaporated toafford the desired product (1.3 g, 6.01 mmol, 57%) as a white solid,LCMS (m/z) 215.1 (M+H)⁺.

Synthesis of phenyl pyrimidin-2-ylcarbamate

To a solution of phenyl carbonochloridate (2.140 g, 13.67 mmol), andpyridine (1.361 mL, 16.82 mmol) in dichloromethane (DCM) (10 mL) stirredunder nitrogen at room temperature was added pyrimidin-2-amine (1.0 g,10.51 mmol). The reaction mixture was stirred at room temperature for 30min. The reaction mixture was quenched with saturated sodium bicarbonatesolution. Separated organic layer and the aqueous layer extracted withDCM (50 ml). Combined DCM layer washed with water and dried out withsodium sulfate, filtered and concentrated under high vacuum to get crudeproduct. This was added to a silica gel column and was eluted with 20%EtOAc/Hexane. Collected fractions were evaporated to afford the desiredproduct (1.6 g, 6.49 mmol, 61.7%), LCMS (m/z) 216.3 (M+H)⁺.

Synthesis of phenyl (5-fluoropyridin-2-yl)carbamate

To a solution of phenyl carbonochloridate (1.397 g, 8.92 mmol), andPyridine (0.721 mL, 8.92 mmol) in dichloromethane (DCM) (40 mL) stirredunder nitrogen at room temp was added 5-fluoropyridin-2-amine (1.0 g,8.92 mmol). The reaction mixture was stirred at RT for 30 min. Thereaction mixture was quenched with saturated sodium bicarbonatesolution. Separated organic layer and the aqueous layer extracted withDCM (20 ml). Combined organic layer washed with water followed by brinesolution and dried out with sodium sulfate, filtered and concentratedunder vacuum to give the desired product (1.4 g, 5.94 mmol, 67%), LCMS(m/z) 233.2 (M+H)⁺.

Synthesis of phenyl (2-methyl-2H-indazol-5-yl)carbamate

To a solution of phenyl carbonochloridate (1.064 g, 6.79 mmol), andpyridine (0.550 mL, 6.79 mmol) in Dichloromethane (DCM) (40 mL) stirredunder nitrogen at room temp was added 2-methyl-2H-indazol-5-amine (1 g,6.79 mmol). The reaction mixture was stirred at room temperature for 30min. The reaction mixture was quenched with saturated sodium bicarbonatesolution. Separated organic layer, aqueous layer extracted with DCM (20ml). Combined organic layer washed with water followed by brine solutionand dried out with sodium sulfate and concentrated under vacuum to getphenyl (2-methyl-2H-indazol-5-yl)carbamate (1.3 g, 4.82 mmol, 70.9%yield), LCMS (m/z) 268.1 (M+H)⁺.

Synthesis of phenyl pyridazin-3-ylcarbamate

To a solution of phenyl carbonochloridate (1.070 g, 6.83 mmol), pyridine(0.665 g, 8.41 mmol) in dichloromethane (10 ml) stirred under nitrogenat 25° C. was added a suspension of pyridazin-3-amine (0.5 g, 5.26 mmol)in dichloromethane (5 ml) during 5 min. The reaction mixture was stirredat 25° C. for 1 hr. Next, the organic phase was washed with water 3 mL,saturated brine 3 mL, dried over sodium sulfate and concentrated invacuo to give the crude product as a white solid. The compound waswashed with hexane, dried under reduced pressure, LCMS (m/z) 216.2(M+H)⁺.

Synthesis of phenyl pyrimidin-4-ylcarbamate

To a solution of phenyl carbonochloridate (1.070 g, 6.83 mmol), pyridine(0.665 g, 8.41 mmol) in DCM (15 ml) stirred under nitrogen at 25° C. wasadded a suspension of pyrimidin-4-amine (0.5 g, 5.26 mmol) in DCM (5 ml)dropwise during 5 min. The reaction mixture was stirred at 25° C. for 1hr. The organic phase was washed with water 3 mL, brine 3 mL, dried oversodium sulfate and concentrated under vacuo to give the crude product asa off-white solid. The crude compound was washed with Hexane and thendried under reduced pressure to give the desired product (500 mg, 1.95mmol, 37%), LCMS (m/z) 215.9 (M+H)⁺.

Synthesis of 3-((6-aminopyridin-2-yl)oxy)propan-1-ol

propane-1,3-diol (1.358 g, 17.84 mmol) was added to a stirred solutionof NaH (1.070 g, 44.6 mmol) in N-Methyl-2-pyrrolidone (NMP) (5 mL) at 0°C. and stirred for 1 h and followed by addition of6-fluoropyridin-2-amine (1.0 g, 8.92 mmol) and stirred for 2 h at 80° C.Reaction mass was cooled to room temperature, slowly added to ice coldwater and diluted with ethyl acetate. The separated organic layer waswashed with water and brine. The organic layer was dried over Na₂SO₄,filtered and concentrated to obtain crude compound. The crude compoundwas purified by using 100-200 silica gel and eluted in 100% ethylacetate to afford 3-((6-aminopyridin-2-yl)oxy)propan-1-ol (0.4 g, 1.760mmol, 19.73% yield) as brown viscous, LCMS (m/z): 169.22 [M+H]⁺.

Synthesis of 6-(3-((tetrahydro-2H-pyran-2-yl)oxy)propoxy)pyridin-2-amine

3-((tetrahydro-2H-pyran-2-yl)oxy)propan-1-ol (4.2 g, 26.2 mmol) in1,4-Dioxane (20 mL) was added to a solution of NaH (1.307 g, 32.7 mmol)in 1,4-Dioxane (20 mL) at 0° C., and the reaction mixture was stirredfor 30 min at 28° C. 6-chloropyridin-2-amine (2.8 g, 21.78 mmol) in1,4-Dioxane (20 mL) was added to the reaction mixture at 0° C., and thereaction mixture was stirred for 10 hr at 100° C. The reaction mixturewas partitioned between water (20 mL) and DCM (2×25 mL). DCM layer waswashed with saturated NaHCO₃ solution, was separated and dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to crude6-(3-((tetrahydro-2H-pyran-2-yl)oxy)propoxy)pyridin-2-amine (5.5 g,18.12 mmol, 83% yield) as brown oil, LCMS (m/z): 253.2 [M+H]⁺.

Synthesis of 3-((tetrahydro-2H-pyran-2-yl)oxy)propan-1-ol

p-toluenesulfonic acid monohydrate (0.678 g, 3.57 mmol) was added to astirred solution of propane-1,3-diol (5.43 g, 71.3 mmol), and3,4-dihydro-2H-pyran (3 g, 35.7 mmol) in Dichloromethane (DCM) (50 mL)at 0° C. The reaction mixture was stirred for 2 h at 28° C. The reactionmixture was partitioned between water (20 mL) and DCM (2×25 mL). DCMlayer was washed with saturated NaHCO₃ solution, was separated and driedover anhydrous Na₂SO₄, filtered and filtrate was evaporated to crude3-((tetrahydro-2H-pyran-2-yl)oxy)propan-1-ol (4.2 g, 26.2 mmol, 73.5%yield) as colorless oil.

Synthesis of(S)-3-bromo-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridine

Cesium carbonate (37.0 g, 114 mmol) was taken into multi-neck RB. Thenflask was cooled to 0° C. and N-Methyl-2-pyrrolidone (NMP) (100 mL) wasadded slowly over a period of 3 minutes. The resulting reaction mixturewas stirred under nitrogen for 15 min. Then(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (10 g, 76 mmol) was addeddropwise over a period of 5 min at 0° C. This suspension was stirred atroom temperature ° C. for 1 h. Suspension became pale yellow solutionafter added 3-bromo-5-fluoropyridine (7.62 mL, 73.9 mmol). The resultingsolution was stirred at 75° C. for 24 hr. Reaction progress wasmonitored by TLC 40% EtOAc in Hexane. TLC indicated consumption of SMand formation of new spot after 24 h. The reaction mass was cooled toroom temperature, diluted with water (500 mL). The aqueous layer wasextracted with ethyl acetate (2×300 mL). The organic layer was washedwith brine (250 mL), dried over Na₂SO₄ filtered, concentrated underreduced pressure to afford brown oil. The crude product was purified bycolumn chromatography over 100-200 mesh size silica gel. Column waseluted with a gradient of EtOAc/Hexane. Desired compound was eluted with20% EtOAc in Hexane. Compound fractions containing pure compound wereconcentrated under reduced pressure to afford(S)-3-bromo-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridine (10 g,34.0 mmol, 44.9% yield) as pale yellow viscous oil, LCMS (m/z): 289.99[M+H]⁺.

Synthesis of(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine

(R)-3-bromo-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridine (50 g,174 mmol), liquor ammonia (25 mL, 1155 mmol) were taken in a sealedtube. Then added copper(II) sulfate (5.54 g, 34.7 mmol) at 0° C. Theresulting blue solution was heated to 120° C. for 2 hr. The reactionprogress was monitored by TLC 10% MeOH in DCM, TLC indicated formationof new spot and consumption of SM after 24 h. After completion, Thereaction mass was cooled to room temperature. The reaction mass wasbrought to pH 10 with 20% NaOH, saturated with NaCl, extracted withethyl acetate (30 mL*2). The combined organic layer was washed withbrine (20 mL), dried over Na₂SO₄, filtered and concentrated underreduced pressure to afford crude brown solid, which was triturated withdiethyl ether and stirred for 4 hours then filtered to afford(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (35.4 g,146 mmol, 84% yield) as pale brown solid, LCMS (m/z): 225.29 [M+H]⁺.

Synthesis of(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine

(S)-3-bromo-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridine (10 g,34.7 mmol), liquor ammonia (100 mL, 4621 mmol) were taken in a sealedtube. The resulting brown solution was heated to 120° C. for 24 hr. Thereaction progress was monitored by TLC 10% MeOH in DCM, TLC indicatedformation of new spot and consumption of SM after 24 h. Aftercompletion, The reaction mass was cooled to room temperature. Thereaction mass was brought to pH 10 with 20% NaOH, saturated with NaCl,extracted with ethyl acetate (30 mL*2). The combined organic layer waswashed with brine (20 mL), dried over Na₂SO₄, filtered and concentratedunder reduced pressure to afford the(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (6 g,25.8 mmol, 74.2% yield) as an pale brown solid, LCMS (m/z): 225.10[M+H]⁺.

Synthesis of(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine

To a suspension of NaH (11.35 g, 473 mmol) in THF (100 mL) was addeddropwise a solution of (R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (25g, 189 mmol) in THF (150 mL) under Nitrogen at 0° C. The resultingsuspension was stirred at rt for 1 h. 6-chloropyrimidin-4-amine (19.61g, 151 mmol) was added to the reaction mixture portion wise at rt andthe resulting suspension was heated to 90° C. for 48 hr. After thecompletion of reaction (monitored by TLC, it shows little bit ofstarting and new spot observed at polar), reaction mixture was pouredinto ice water (500 mL) and aqueous layer was extracted with EtOAc(2×1000 mL). Combined organics dried over Na₂SO₄, filtered andconcentrated under reduced pressure to get light brown solid (crude).Crude material was purified by silica gel column (100-200, 3% MeOH inDCM). Fractions containing pure compound were combined and concentratedto afford the desired product(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine (13 g,53.9 mmol, 28.5% yield) as an off-white solid and also get the impurecompound (10 g). LCMS (m/z): 226.17 (M+H)⁺.

Synthesis of(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine

To a suspension of NaH (9.08 g, 378 mmol) in THF (150 mL) was added dropwise a solution of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (20 g,151 mmol) in THF (200 mL) under Nitrogen at 0° C., and the resultingsuspension was stirred at rt for 1 h. 6-chloropyrimidin-4-amine (15.68g, 121 mmol) was added to the reaction mass portion wise at rt and theresulting suspension was heated to 90° C. for 48 hr. After thecompletion of reaction (monitored by TLC, starting material completelyconsumed and new spot observed at polar), reaction mass was poured intoice water (200 mL) and extracted with ethyl acetate (2×400 ml). Combinedorganics dried over Na₂SO₄, filtered and concentrated under reducedpressure to get light brown solid. The obtained solid was stirred indiethyl ether (200 ml) for 30 min filtered and dried under vacuum to get(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-amine (13 g,57.3 mmol, 37.9% yield) as a light brown solid, LCMS (m/z): 225.96[M+H]⁺.

Synthesis of(R)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine

To a suspension of NaH (9.08 g, 378 mmol) in THF (150 mL) was addeddropwise a solution of (R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (20g, 151 mmol) in THF (250 mL) under Nitrogen at 0° C. The resultingsuspension was stirred at rt for 1 h. 4-chloropyrimidin-2-amine (15.68g, 121 mmol) was added to the reaction mixture portion wise at rt andthe resulting suspension was heated to 90° C. for 48 hr. After thecompletion of reaction (monitored by TLC, starting completely consumedand new spot observed at polar), reaction mixture was poured into icewater (250 mL) and aqueous layer was extracted with EtOAc (2×300 mL).Combined organics dried over Na₂SO₄, filtered and concentrated underreduced pressure to get pale yellow liquid (crude). Obtained crudematerial was purified by column (100-200 silica gel) by using 0-50%EtOAc-petether to get(R)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (13 g,57.0 mmol, 37.7% yield) as pale yellow solid, LCMS (m/z): 226.20 [M+H]⁺.

Synthesis of(S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine

To a suspension of NaH (8.25 g, 189 mmol) in 1,4-Dioxane (200 mL) wasadded dropwise a solution of(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (10 g, 76 mmol) in1,4-Dioxane (50 mL) under Nitrogen at 0° C. The resulting suspension wasstirred at rt for 1 h. 4-chloropyrimidin-2-amine (7.84 g, 60.5 mmol) wasadded to the reaction mixture portion wise at rt and the resultingsuspension was heated to 90° C. for 48 hr. The reaction mixture wascooled to 28° C. and was partitioned between water (200 mL) and EtOAc(200 mL). Organic layer was separated and was dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to get crude (TLC eluent:Neat ethyl acetate R_(f)0.3; UV active). The crude compound was purifiedby column chromatography (100-200 mesh silica gel, eluted at 60% Ethylacetate in hexane) to afford(S)-4-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (8.0 g,35.4 mmol, 46.8% yield) as pale yellow solid LCMS (m/z) 226.30 (M+H)⁺.

Synthesis of(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine

To a stirred solution of(R)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine (12 g,49.0 mmol) in Tetrahydrofuran (THF) (20 mL) was added ammonium hydroxide(300 mL, 1926 mmol) and copper(II) sulfate (1.566 g, 9.81 mmol) in asealed tube. Reaction mixture was stirred at 120° C. for 18 hr. Progressof the reaction was monitored by TLC, TLC indicates formation of polarspot along with un-reacted SM. Reaction mixture was diluted with water(300 mL), extracted with EtOAc (3×200 mL), organic layers were combinedand washed with water (100 mL), brine solution (100 mL), organic layerdried over anhydrous Na₂SO₄, filtered and concentrated under reducedpressure to afford(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (10 g,3.97 mmol, 8.09% yield) as a yellow oily crude compound, LCMS (m/z):226.13 (M+H)⁺.

Synthesis of(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine

To a suspension of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (8.87 g,67.1 mmol), in N,N-Dimethylformamide (DMF) (50 mL) stirred undernitrogen at 0° C. was added cesium carbonate (32.8 g, 101 mmol), theresulting reaction mixture was stirred at 0° C. for 1 hr. To this added2,5-dichloropyrazine (10 g, 67.1 mmol). The resulting reaction mixturewas stirred at 100° C. for 6 hr. Progress of the reaction was monitoredby TLC. TLC indicated starting material was consumed to form new polarspot with 0.3 Rf. The reaction mass was cooled to rt, added water (100mL) and extracted with Ethyl acetate (100 mL). The organic layer waswashed with water (100 mL×2). The organic layer was dried over Na₂SO₄and filtered and concentrated to get crude as light brown liquid. Thecrude product was added to a silica gel (60-120) column and was elutedwith Hex/EtOAc. Collected fractions: 30% EtOAc in Hexane the product waseluted. Concentrated the product fractions to afford(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine (12 g,47.7 mmol, 71.0% yield) as light brown liquid, LCMS (m/z): 244.90[M+H]⁺.

Synthesis of(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine

To a solution of(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine (10 g,40.9 mmol), in Tetrahydrofuran (THF) (10 mL) stirred at room temp wasadded ammonium hydroxide (63.7 mL, 409 mmol) and copper(II) sulfate(3.26 g, 20.44 mmol) at rt. The reaction mixture was stirred in sealedtube at 130° C. for 2 days. Progress of the reaction was monitored byTLC. TLC indicated starting material was consumed. Cooled the reactionmass to rt, diluted with water (100 mL), Extracted with ethyl acetate(250 mL×2). The organic layer was dried over Na₂SO₄, filtered andconcentrated to get crude compound as brown sticky compound. The crudeproduct was added to a silica gel column and was eluted with DCM/EtOAc.Collected fractions: 50% EtOAc in petether the product was eluted.Concentrated the product fractions to afford(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (2 g,8.77 mmol, 21.46% yield)(N35119-51-A2) as light brown solid. NMR: inCDCl3 consistent with, LCMS (m/z): 226.09 [M+H]⁺.

Synthesis of(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine

(R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (27.8 g, 210 mmol) wasadded to a stirred solution of KOtBu (45.8 g, 408 mmol) in NMP (200 mL)at 0° C. then stirred at RT for 1 h and cooled to 0° C.,6-chloropyridin-3-amine (15 g, 117 mmol) was added and heated to 110° C.for 144 h. The reaction mixture cooled to RT and partitioned betweenwater (500 mL×2) and EtOAc (200 mL×4). Organic layers were separated andwas dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated toget crude and purified by column chromatography (using 100-200 silicagel, column eluted at 50% ethyl acetate in hexane) to afford the(R)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (8 g,35.1 mmol, 30.1% yield) as brown oil, LCMS (m/z): 225.16 [M+H]⁺.

Synthesis of(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine

(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (18.50 g, 140 mmol) wasadded to a stirred solution of KOtBu (30.5 g, 272 mmol) in NMP (600 mL)at 0° C. then stirred at RT for 1 h and cooled to 0° C.,6-chloropyridin-3-amine (10.0 g, 78 mmol) was added and heated to 110°C. for 88 h. The reaction mixture cooled to RT and partitioned betweenwater (50 mL×2) and EtOAc (100 mL×2). Organic layers were separated andwas dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated toget crude compound as a gum. (TLC: Eluent: 100% ethyl acetate, R_(f)0.5;UV active:). The crude product was purified by flash columnchromatography (silica-gel: 100-200 mesh) eluted with 50% EtOAc inhexane to afford(S)-6-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-amine (10.0 g,41.7 mmol, 53.6% yield) as a dark sticky mass, LCMS (m/z) 225.0 (M+H)⁺.

Synthesis of(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine

(R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (30.7 g, 232 mmol) wasadded to a stirred solution of KOtBu (70.1 g, 624 mmol) in NMP (800 mL)at 0° C. then stirred at RT for 1 h and cooled to 0° C. then5-fluoropyridin-2-amine (20 g, 178 mmol) was added and heated to 110° C.for 114 h. The reaction mixture cooled to RT and partitioned betweenwater (500 mL×2) and EtOAc (500 mL×4). Organic layers were separated andwas dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated toget crude compound, then it was purified by column chromatography (using100-200 silica gel, column eluted at 80% ethyl acetate in hexane) toafford the(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (10 g,40.1 mmol, 22.50% yield) as a brown oil, LCMS: 225.0 (M+H).

Synthesis of(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine

NaH (12.84 g, 268 mmol) was added to a stirred solution of(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (31.8 g, 241 mmol) inDimethyl Sulfoxide (DMSO) (100 mL) at 0° C. then stirred at RT for 1 hand cooled to 0° C., 5-fluoropyridin-2-amine (15.0 g, 134 mmol) wasadded and heated to 110° C. for 60 h. The reaction mixture cooled to RTand partitioned between water (50 mL) and EtOAc (100 mL). Organic layerswere separated and was dried over anhydrous Na₂SO₄, filtered andfiltrate was evaporated to get crude compound (TLC: Eluent: 100% ethylacetate, R_(f)0.5; UV active), The crude product was purified by flashcolumn chromatography (silica-gel: 100-200 mesh) eluted with 50% EtOAcin hexane to afford(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-amine (7.2 g,32.1 mmol, 23.99% yield) as a pale yellow sticky, LCMS (m/z): 225.1(M+H)⁺.

Synthesis of(R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-amine

Tetrahydrofuran (75 mL) was added to NaH (5.56 g, 232 mmol) at 0° C.,(R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (12.46 mL, 100 mmol) inTetrahydrofuran (50 mL was added to the reaction mixture at 0° C., andthe reaction mixture was stirred for 1 h at 28° C.2-chloropyrimidin-5-amine (10 g, 77 mmol) in Tetrahydrofuran (25 mL) wasadded and stirred for 16 hr at 70° C. The reaction mixture was quenchedwith cold water (30 mL) and extracted with ethyl acetate (3×80 mL). Theorganic layer was washed with water (2×50 mL) and saturated brinesolution (50 mL), dried over anhydrous Na₂SO₄, filtered andconcentrated. The crude compound was purified by column chromatography(Neutral alumina) product was eluted with 40-45% Ethyl acetate in Hexaneto afford(R)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-amine (6.5 g,28.3 mmol, 36.6% yield) as pale yellow solid, LCMS (m/z): 226.0 [M+H]⁺.

Synthesis of(S)-2-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-amine

To a suspension of NaH (6.17 g, 154 mmol) in THF (100 ml) was added(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol (13.26 g, 100 mmol) in THF(50 ml) was added to the reaction mixture at 0° C., and the reactionmixture was stirred for 1 h at 25° C. to this 2-chloropyrimidin-5-amine(10 g, 77 mmol) in THF (50 ml) and was added at 0° C. and slowly heatedto 80° C. and stirred for 16 hr at 80° C. After completion of thereaction, reaction mixture was quenched with the ammonium chloride (10ml) and extracted with the ethyl acetate (3×20 ml). The organic layerwas separated and washed with the brine and dried over Na₂SO₄, filteredit and concentrated under reduced pressure to get the crude. This crudewas triturated with the diethyl ether to get(S)-2-(2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-amine (5.0 g,19.77 mmol, 25.6% yield) as a brown solid, LCMS (m/z): 226.1 [M+H]⁺.

Synthesis of (S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl) methyl)pyrazine

To a stirred solution of cesium carbonate (492 g, 1510 mmol) in DMF(1000 mL) was added (S)-(2,2-dimethyl-1,3-dioxolan-4-yl) methanol (133g, 1007 mmol) at 0° C. The resulting reaction mixture was stirred atroom temperature for 30 min. Then a solution of 2,5-dichloropyrazine(150 g, 1007 mmol) in DMF (500 mL) was added at 0° C. and the resultedreaction mixture was stirred at 100° C. for 4 h. (TLC System: 20% Ethylacetate in Petether, R_(f): 0.5, UV active). The reaction mixture wasdiluted with ice cold water (500 mL), extracted with EtOAc (3×300 mL).The combined organic layer was washed with water (2×200 mL) and brinesolution (100 mL), dried over anhydrous Na₂SO₄, filtered andconcentrated under reduced pressure to obtain crude compound. The crudecompound was purified by flash column chromatography (silica gel:100-200 mesh, eluent: 10% EtOAc in Hexane) to afford the desired product(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl) methoxy) pyrazine (200g, 768 mmol, 76% yield) as a yellow liquid. LCMS (m/z): 245.1 [M+H]⁺.

Synthesis of(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine

To a stirred solution of(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine (120 g,490 mmol) in THF (30 mL) were added ammonium hydroxide (1000 mL, 6420mmol) and copper(II) sulfate (15.66 g, 98 mmol) in a sealed tube and theresult reaction mixture was stirred at 120° C. for 48 h (TLC System: 50%Ethyl acetate in Petether, R_(f) 0.4, UV active). The reaction mixturewas diluted with water (300 mL), extracted with EtOAc (3×500 mL). Thecombined organic layer was washed with water (200 mL) and brine solution(200 mL), dried over anhydrous Na₂SO₄, filtered and concentrated underreduced pressure to get crude compound. The crude was purified by flashcolumn chromatography (using 100-200 mesh silicagel and eluted thecompound with 40% EtOAc in Hexane) to afford the desired product(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (65 g,280 mmol, 57.2% yield) as a yellow crystal solid. LCMS (m/z): 226.13[M+H]⁺.

Synthesis of (R)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine

To a stirred suspension of cesium carbonate (32.8 g, 101 mmol) in DMF(100 mL) was added (R)-(2,2-dimethyl-1,3-dioxolan-4-yl) methanol (8.87g, 67.1 mmol) at 0° C. and stirred at room temperature for 30 min. Then2,5-dichloropyrazine (10 g, 67.1 mmol) was added and the resultingreaction mixture was stirred at 100° C. for 4 h. (TLC System: 20% Ethylacetate in Hexane, R_(f): 0.5, UV active). The reaction mixture wasdiluted with ice cold water (200 mL), extracted with EtOAc (3×100 mL).The combined organic layer was washed with water (2×50 mL) and brinesolution (50 mL), dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure to afford(R)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine (12 g,43.8 mmol, 65.3% yield) as a yellow oily compound. LCMS (m/z): 244.99[M+H]⁺.

Synthesis of (R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl) methoxy)pyrazin-2-amine

To a stirred solution of(R)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazine (8 g,32.7 mmol) in Tetrahydrofuran (10 mL) was added ammonium hydroxide (400mL, 2568 mmol) and copper(II) sulfate (1.044 g, 6.54 mmol) in a sealedtube and the reaction mixture was stirred at 120° C. for 48 h. (TLCSystem: 50% Ethyl acetate in Hexane, R_(f) 0.4, UV active). The reactionmixture was diluted with water (200 mL), extracted with EtOAc (3×50 mL).The combined organic layer was washed with water (50 mL), brine solution(50 mL), dried over anhydrous Na₂SO₄, filtered and concentrated underreduced pressure to get crude compound. The crude was purified by flashcolumn chromatography (using 100-200 mesh silicagel and eluted thecompound with 40% EtOAc in Hexane), pure fraction were collected andconcentrated under reduced pressure to afford(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-amine (2 g,8.65 mmol, 26.4% yield) as a yellow crystal solid. LCMS (m/z): 226.10[M+H]⁺.

Synthesis of(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidine

To a stirred solution of 2-chloropyrimidin-5-ol (13 g, 100 mmol) in THF(100 mL) at 0° C. was added (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol(13.16 g, 100 mmol), triphenylphosphine (32.7 g, 124 mmol) followed byDEAD (19.71 mL, 124 mmol) and reaction was stirred at RT for 4 h. (TLCeluting system: 30% EtOAc in pet ether; R_(f)-0.5; UV active). Thereaction mixture was quenched with water (50 mL) and extracted intoEtOAc (2×75 mL). Organic layer was separated and dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to give crude product. Thecrude was purified by chromatography (Silicagel, eluent: 20% EtOAc inhexane) to afford(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidine (20g, 79 mmol, 79% yield) as an off white solid. LCMS (m/z): 245.10;[M+H]⁺.

Synthesis of(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine

A mixture of(S)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidine (10g, 40.9 mmol) and aq.ammonia (66.3 ml, 1226 mmol) in a sealed tube washeated at 120° C. for 24 h. (TLC eluting system: 100% EtOAc; R_(f)-0.2;UV active). The reaction mixture was cooled to RT, quenched with water(50 mL) and extracted into EtOAc (2×75 mL). Organic layer was separated,dried over anhydrous sodiumsulphate, filtered and filtrate wasevaporated to give crude product as yellow solid. The crude compound wastriturated with n-pentane (50 mL) to afford(S)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (6.6 g,28.6 mmol, 70.0% yield) as an off white solid. LCMS (m/z): 226.17;[M+H]⁺.

Synthesis of(R)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidine

To a stirred solution of 2-chloropyrimidin-5-ol (20 g, 153 mmol) in THF(100 mL) at 0° C. was added (R)-(2,2-dimethyl-1,3-dioxolan-4-yl)methanol(24.30 g, 184 mmol), triphenylphosphine (50.2 g, 192 mmol) followed byDEAD (30.3 mL, 192 mmol) and the reaction was stirred at RT for 12 h.(TLC eluting system: 70% EtOAc in pet ether; R₁-0.5; UV active). Thereaction mixture was quenched with water (100 mL) and extracted intoEtOAc (200 mL). Organic layer was separated and dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to give crude product. Thecrude was purified by chromatography (Silicagel, eluent: 35% EtOAc inhexane) to afford(R)-2-chloro-5-(2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidine (23 g,91 mmol, 59.5% yield) as a white solid. LCMS (m/z): 245.06; [M+H]⁺.

Synthesis of(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine

A mixture of(R)-2-chloro-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidine (5 g,20.44 mmol) and aq.ammonia (50 ml, 924 mmol) in a sealed tube was heated120° C. for 48 h. (TLC eluting system: 100% EtOAc; R_(f)-0.2; UVactive). The reaction mixture was cooled to RT, quenched with water (50mL) and extracted into DCM (2×75 mL). Organic layer was separated, driedover anhydrous Na₂SO₄, filtered and filtrate was evaporated to afford(R)-5-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-amine (2.7 g,11.5 mmol, 57.5% yield) as a pale yellow solid. LCMS (m/z): 226.02;[M+H]⁺.

Compound Examples Example 1 Synthesis of(9S)-N-(pyridin-2-yl)-2-((S)-2-(trifluoromethyl)pyrrolidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

NaH (0.254 g, 10.57 mmol was added to a stirred solution of(9S)-2-((S)-2-(trifluoromethyl)pyrrolidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.550 g, 1.761 mmol) in Tetrahydrofuran (THF) (30 mL) stirred undernitrogen at room temp. The reaction mixture was stirred at room temp for30 minutes. Then3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (0.635 g,2.64 mmol) was added at room temp. Then reaction mixture was stirred at65° C. for 24 hr. The reaction mixture was cooled to 28° C. and waspartitioned between water (10 mL) and EtOAc (25 mL). Organic layer wasseparated and was dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to give crude as brown solid (TLC eluent: 100% EtOAc:R_(f)-0.3; UV active). The crude was purified by column chromatographyusing (100-200 mesh) silica gel and was eluted with 75-80% EtOAc inHexane to afford pure(9S)-N-(pyridin-2-yl)-2-((S)-2-(trifluoromethyl)pyrrolidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.275 g, 0.631 mmol, 35.8% yield) as off-white solid, LCMS (m/z): 433.2[M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.60 (s, 1H), 8.20-8.29 (m, 1H), 8.09(dt, J=8.33, 0.88 Hz, 1H), 7.72-7.81 (m, 1H), 7.37 (d, J=8.55 Hz, 1H),7.05 (ddd, J=7.29, 4.88, 0.99 Hz, 1H), 6.45 (d, J=8.55 Hz, 1H),5.11-5.19 (m, 1H), 4.74 (br s, 1H), 3.91 (dt, J=10.19, 5.21 Hz, 1H),3.50-3.62 (m, 1H), 3.27 (d, J=1.75 Hz, 1H), 3.11-3.22 (m, 1H), 3.01-3.08(m, 1H), 2.81 (br d, J=13.37 Hz, 1H), 2.07-2.28 (m, 4H), 1.94-2.02 (m,1H), 1.81-1.92 (m, 1H), 1.21-1.40 (m, 2H)

Example 2 Synthesis of(9S)-N-(5-fluoropyridin-3-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.561 mmol) in THF (15 mL) were added triethylamine (0.653 mL,4.68 mmol) and triphosgene (232 mg, 0.780 mmol) at 30° C. and stirred atroom temperature for 1 h. Then 5-fluoropyridin-3-amine (525 mg, 4.68mmol) was added at 30° C. and reaction was heated at 70° C. for 16 h.The THF evaporated under reduced pressure and the obtained residue wasdiluted with water and extracted with CH₂Cl₂ (3×50 mL). The combinedorganic layer was washed with water, brine, dried over Na₂SO₄ andsolvent was evaporated under reduced pressure to obtain the crudecompound and it was purified by flash column chromatography (silica-gel:100-200 mesh, eluent: 3% MeOH in DCM) to afford(9S)-N-(5-fluoropyridin-3-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.4366 mmol, 35% yield) as an off-white solid (TLC: R_(f)=0.3,10% MeOH in EtOAc), LCMS (m/z): 459.25[M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.19 (s, 1H), 8.93 (d, J=5.04 Hz, 1H),8.47 (s, 1H), 8.37 (s, 1H), 8.33-8.21 (m, 2H), 8.03 (d, J=11.18 Hz, 1H),7.88 (d, J=8.11 Hz, 1H), 7.78 (d, J=8.11 Hz, 1H), 4.83 (s, 1H), 3.43 (d,J=12.06 Hz, 1H), 3.25 (s, 2H), 2.90 (d, J=13.81 Hz, 1H), 2.11-1.84 (m,2H), 1.33 (s, 2H).

Example 3 Synthesis of(9S)-N-(pyrimidin-5-yl)-2-(5-(trifluoromethyl)pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(5-(trifluoromethyl)pyridin-3-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.6 g, 1.873 mmol), triethylamine (0.783 mL, 5.62 mmol) in THF (12 mL)was added triphosgene (0.278 g, 0.937 mmol) at 0° C. and stirred at RTfor 30 min. Then Pyrimidine-5-amine was added and the reaction mixturewas stirred at 65° C. for 16 h. The solvent was removed under reducedpressure to obtain the crude and diluted with dichloromethane, washedwith water and brine solution and dried over anhydrous Na₂SO₄,evaporated the organic layer under reduced pressure to obtain the crudeproduct. The crude mixture was purified by flash column chromatography(silica-gel: 100-200 mesh, eluted with 2% MeOH in CH₂Cl₂) to afford(9S)-N-(pyrimidin-5-yl)-2-(5-(trifluoromethyl)pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(459 mg, 1.016 mmol, 54.2% yield) as a white solid (TLC: 80% EtOAc inHexane, R_(f)=0.5), LCMS (m/z): 442.2 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.19 (s, 1H), 9.48 (d, J=2.19 Hz, 1H),9.08 (s, 1H), 8.96 (s, 1H), 8.88 (s, 1H), 8.71 (s, 1H), 7.85 (d, J=8.11Hz, 1H), 7.77 (d, J=7.89 Hz, 1H), 4.84 (s, 1H), 3.43 (dd, J=13.59, 1.97Hz, 1H), 3.33 (s, 2H), 2.90 (d, J=13.81 Hz, 2H), 1.95 (d, J=10.30 Hz,2H), 1.33 (m, 2H).

Example 4 Synthesis of(9S)-2-(2-methylpyridin-4-yl)-N-(pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

NaH (0.297 g, 12.39 mmol) was added to a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.550 g, 2.065 mmol) in Tetrahydrofuran (THF) (30 mL) stirred undernitrogen at room temperature. Then the reaction mixture was stirred atroom temperature for 30 minutes.3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (0.744 g,3.10 mmol) was added at was added at room temperature. Then the reactionmixture was stirred at 65° C. for 24 hr. The reaction mixture was cooledto 28° C. and was partitioned between water (20 mL) and EtOAc (50 mL).Organic layer was separated and was dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to give crude as brown solid (TLCeluent: 70% EtOAc in Hexane: R_(f)=0.3; UV active). The crude waspurified by column chromatography using (100-200 mesh) silica gel andwas eluted with 50-60% EtOAc in Hexane to afford pure(9S)-2-(2-methylpyridin-4-yl)-N-(pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.292 g, 0.751 mmol, 36.4% yield) as a pale yellow solid, LCMS (m/z):387 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.99 (s, 1H), 8.59 (d, J=5.48 Hz, 1H),8.41-8.45 (m, 1H), 8.27 (s, 1H), 8.18 (d, J=8.33 Hz, 1H), 7.97 (br d,J=3.73 Hz, 1H), 7.82-7.86 (m, 2H), 7.71 (d, J=7.89 Hz, 1H), 7.11-7.16(m, 1H), 4.86 (br s, 1H), 3.40 (br d, J=13.37 Hz, 1H), 3.29 (s, 2H),2.91 (br d, J=13.15 Hz, 1H), 2.64 (s, 3H), 1.93-2.05 (m, 2H), 1.32 (brs, 2H)

Example 5 Synthesis of(9S)-N-(5-fluoropyridin-3-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.6 g, 2.253 mmol) in Tetrahydrofuran (THF) (30 mL) stirred undernitrogen at room temperature was added triethylamine (1.884 mL, 13.52mmol) and triphosgene (0.669 g, 2.253 mmol). Then reaction mixture wasstirred at rt for 30 minutes. Next, 5-fluoropyridin-3-amine (0.758 g,6.76 mmol) was added at rt. and the reaction mixture was stirred at 65°C. for 16 hr. The reaction mixture was cooled to room temp. The reactionmixture concentrated under reduced pressure and then partitioned betweenwater (20 mL) and Dichloromethane (50 mL). Organic layer was separatedand was dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to give crude as brown solid (TLC eluent: 100% EtOAc:R_(f)-0.4; UV active). The crude was purified by column chromatographyusing (100-200 mesh) silica gel and was eluted with 100% EtOAc to affordpure(9S)-N-(5-fluoropyridin-3-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.314 g, 0.770 mmol, 34.2% yield) as a pale yellow solid, LCMS (m/z):405.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ 13.71 (s, 1H), 8.74-8.55 (m, 1H), 8.44 (d,J=1.7 Hz, 1H), 8.29 (d, J=2.6 Hz, 1H), 8.18-7.99 (m, 1H), 7.81 (d, J=1.7Hz, 1H), 7.73 (d, J=2.8 Hz, 3H), 4.82 (s, 1H), 3.39 (d, J=1.9 Hz, 1H),3.30 (s, 2H), 2.92 (s, 1H), 2.57 (s, 3H), 2.01 (s, 2H), 1.32 (s, 2H).

Example 6 Synthesis of(9S)-N-(pyridazin-3-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.4 g, 1.249 mmol), phenyl pyridazin-3-ylcarbamate (0.806 g, 3.75 mmol)in THF (8 mL) was added DMAP (0.458 g, 3.75 mmol) at 25° C. undernitrogen. The reaction mixture was heated at 80° C. for 16 h. Allowed tocool to RT and the solvent was removed in vacuo to obtain crude wasdiluted with CH₂Cl₂ and washed with water, brine solution, dried overanhydrous Na₂SO₄. The organic layer was concentrated in vacuo to obtainthe crude compound. The crude mixture was purified by flash columnchromatography (silica-gel: 100-200 mesh, eluent: 4% MeOH in CH₂Cl₂) toafford(9S)-N-(pyridazin-3-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(320 mg, 0.711 mmol, 56.9% yield) as a white solid (TLC: 80% EtOAc inHexane R_(f): 0.6), LCMS (m/z): 442.2 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d6): δ 14.27 (s, 1H), 8.98 (dd, J=4.7, 1.5 Hz,1H), 8.90 (d, J=5.1 Hz, 1H), 8.59-8.48 (m, 2H), 8.38 (dd, J=9.0, 1.5 Hz,1H), 8.01 (d, J=8.1 Hz, 1H), 7.79 (d, J=8.1 Hz, 1H), 7.71 (dd, J=9.0,4.7 Hz, 1H), 5.25-4.49 (m, 1H), 3.42 (dd, J=13.7, 1.9 Hz, 1H), 3.30 (s,2H), 2.94 (dt, J=13.8, 2.5 Hz, 1H), 2.02 (m, 2H), 1.34 (q, J=5.2, 4.4Hz, 2H).

Example 7 Synthesis of(9S)-N-(pyridin-2-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.250 g, 0.780 mmol) in THF (5 mL) was added sodium hydride (0.156 g,3.90 mmol) at RT in one charge. The reaction mixture was stirred at RTfor 30 min. Then3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (0.562 g,2.341 mmol) was added and stirred the reaction mixture at 65° C. for 16h. Allowed to cool to room temperature and quenched it with ice coldwater, extracted with ethyl acetate (3×50 mL). The combined organiclayer was washed with water, saturated brine solution, dried over sodiumsulfate and concentrated under reduced pressure to obtain crude product.The crude mixture was purified by flash column chromatography(silica-gel: 100-200 mesh, eluent: 2% MeOH in CH₂Cl₂) to afford(9S)-N-(pyridin-2-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (302 mg, 0.681 mmol, 87%yield) as a white solid (TLC: 70% EtOAc in Hexane, R_(f): 0.5), LCMS(m/z): 441.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d6): δ 13.85 (s, 1H), 8.94 (d, J=5.2 Hz, 1H), 8.64(dd, J=1.7, 0.9 Hz, 1H), 8.55-8.50 (m, 1H), 8.34 (ddd, J=4.8, 2.0, 0.9Hz, 1H), 8.15 (dt, J=8.3, 1.0 Hz, 1H), 7.98 (d, J=8.1 Hz, 1H), 7.86-7.79(m, 1H), 7.77 (d, J=8.0 Hz, 1H), 7.12 (ddd, J=7.3, 4.9, 1.0 Hz, 1H),4.87 (t, J=2.6 Hz, 1H), 3.41 (dd, J=13.6, 1.9 Hz, 1H), 3.30 (s, 2H),2.92 (d, J=14.0 Hz, 1H), 2.09-1.86 (m, 2H), 1.34 (d, J=8.5 Hz, 2H).

Example 8 Synthesis of(9S)-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

NaH (0.277 g, 11.53 mmol) was added to a solution of(9S)-2-(3)pyrrolidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-metha-(trifluoromethylnopyrido[2,3-b][1,4]diazocine (0.6 g, 1.921 mmol) in Tetrahydrofuran(THF) (20 mL) and stirred for 30 min. Then3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (0.692 g,2.88 mmol) was added at room temp. Then reaction mixture was stirred at65° C. for 24 h. The reaction mixture was cooled to RT and waspartitioned between water (20 mL) and EtOAc (50 mL). Organic layer wasseparated and was dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to give crude as brown solid (TLC eluent: 70% EtOAc inHexane: R_(f)-0.3; UV active). The crude was purified by columnchromatography using (100-200 mesh) silica gel and was eluted with50-60% EtOAc in hexane to afford mixture of stereoisomers (9:1) which onfurther SFC purification obtained pure(9S)-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.422 g, 0.973 mmol, 50.6% yield) as an off-white solid, LCMS (m/z):433.3 [M+H]⁺.

Analytical SFC Conditions: Peak-I Column/dimensions: Chiralcel OX-H(250×4.6) mm, 5 u % CO₂: 60.0%

% Co solvent: 40.0% (0.5% DEA IN MeOH)Total Flow: 4.0 g/min

Back Pressure: 100 bar Temperature: 26.8° C. UV: 263 nm

¹H NMR (400 MHz, DMSO-d₆): δ 13.81 (s, 1H), 8.23-8.18 (m, 1H), 8.11 (dt,J=8.3, 1.0 Hz, 1H), 7.76 (ddd, J=8.5, 7.4, 1.9 Hz, 1H), 7.32 (d, J=8.5Hz, 1H), 7.04 (ddd, J=7.3, 4.9, 1.0 Hz, 1H), 6.24 (d, J=8.5 Hz, 1H),4.71 (s, 1H), 3.96 (d, J=8.5 Hz, 1H), 3.78 (d, J=7.0 Hz, 1H), 3.74 (s,1H), 3.66 (d, J=4.1 Hz, 2H), 3.59-3.38 (m, 1H), 3.26 (d, J=1.9 Hz, 1H),3.12 (dd, J=12.7, 3.5 Hz, 1H), 3.05 (s, 1H), 2.80 (d, J=13.4 Hz, 1H),2.41-2.29 (m, 1H), 2.15 (dd, J=12.7, 8.1 Hz, 1H), 1.96 (s, 1H), 1.83 (s,2H), 1.31 (s, 2H).

Example 9 Synthesis of(9S)-N-(pyrimidin-4-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

(9S)-2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 1.561 mmol), triethylamine (0.653 mL, 4.68 mmol) in THF (10 mL)stirred under nitrogen at 25° C. was added bis(trichloromethyl)carbonate (0.232 g, 0.780 mmol) in one charge. The reaction mixture wasstirred at 25° C. for 30 min. Then pyrimidin-4-amine (0.445 g, 4.68mmol) was added in a portion, reaction was stirred at 65° C. for 16 h.Allowed to cool to RT and solvent was removed under vacuo, crude wasdiluted with DCM, washed with water, brine solution, dried over sodiumsulfate and concentrated under vacuum to obtain crude compound. Thecrude mixture was purified by prep HPLC to afford(9S)-N-(pyrimidin-4-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(199 mg, 0.446 mmol, 28.6% yield) as a white solid (TLC: 70% EtOAc inHexane, R_(f): 0.6), LCMS (m/z): 442.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d6): δ 14.18 (s, 1H), 8.97 (d, J=5.1 Hz, 1H), 8.85(d, J=1.2 Hz, 1H), 8.69 (d, J=5.8 Hz, 1H), 8.60 (dd, J=1.9, 0.8 Hz, 1H),8.50 (dd, J=5.1, 1.7 Hz, 1H), 8.12 (dd, J=5.8, 1.3 Hz, 1H), 8.02 (d,J=8.1 Hz, 1H), 7.80 (d, J=8.1 Hz, 1H), 4.93-4.73 (m, 1H), 3.42 (dd,J=13.7, 1.9 Hz, 1H), 3.32 (d, J=8.2 Hz, 2H), 2.93 (dt, J=13.9, 2.5 Hz,1H), 2.12-1.83 (m, 2H), 1.33 (d, J=7.8 Hz, 2H).

Example 10 Synthesis of(9S)-N-(pyrimidin-5-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 1.561 mmol), triethylamine (0.653 mL, 4.68 mmol) in THF (10 mL)was added bis(trichloromethyl) carbonate (0.232 g, 0.780 mmol) at 25° C.under nitrogen atmosphere and stirred at RT for 30 min. Thenpyrimidin-5-amine (0.445 g, 4.68 mmol) was added and heated the reactionmixture at 65° C. for 16 h. Allowed to cool to RT and solvent wasremoved on rota-vapour, the crude was diluted with CH₂Cl₂ (20 ml) andwashed with water (5 ml), brine solution (5 ml) followed by dried oversodium sulfate. The organic solvent was evaporated under reducedpressure to obtain the crude product. The crude mixture was purified byprep-HPLC (formic acid, ACN 25%) to afford(9S)-N-(pyrimidin-5-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.56 mmol, 45% yield) as an off white solid (TLC: 80% EtOAc inHexane, R_(f): 0.5), LCMS (m/z): 442.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d6): δ 13.07 (s, 1H), 9.08-8.72 (m, 4H), 8.38 (dd,J=1.7, 0.9 Hz, 1H), 8.28 (dd, J=5.2, 1.6 Hz, 1H), 7.90 (d, J=8.0 Hz,1H), 7.79 (d, J=8.0 Hz, 1H), 4.85 (t, J=2.7 Hz, 1H), 3.43 (dd, J=13.7,1.9 Hz, 1H), 3.33 (d, J=10.1 Hz, 2H), 2.91 (d, J=13.7 Hz, 1H), 2.15-1.79(m, 2H), 1.33 (s, 2H).

Example 11 Synthesis of(9S)-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

NaH (0.110 g, 4.60 mmol) was added to a stirred solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-II from the intermediate SFC separation, 0.250 g, 0.766 mmol) inTetrahydrofuran (THF) (20 mL) stirred under nitrogen at room temp. Thereaction mixture was stirred at room temp for 30 minutes. Then3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (0.276 g,1.149 mmol) was added at room temp° C. Then reaction mixture was stirredat 65° C. for 24 hr. Reaction was cooled to room temperature and waspartitioned between water (10 mL) and EtOAc (50 mL). Organic layer wasseparated and was dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to give crude as brown solid (TLC eluent: 70% EtOAc inHexane: R_(f)-0.3; UV active). The crude was purified by columnchromatography using (100-200 mesh) silica gel and was eluted with50-60% EtOAc in Hexane to afford pure afford(9S)-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.2 g, 0.439 mmol, 57.3% yield) as a off-white solid, LCMS (m/z): 447.2[M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ 13.50 (s, 1H), 8.32-8.18 (m, 1H), 8.16-8.03(m, 1H), 7.76 (ddd, J=8.8, 7.4, 2.0 Hz, 1H), 7.35 (d, J=8.6 Hz, 1H),7.05 (dd, J=7.8, 4.5 Hz, 1H), 6.60 (d, J=8.7 Hz, 1H), 4.72 (d, J=2.7 Hz,1H), 4.27 (s, 2H), 3.31-3.24 (m, 1H), 3.22-2.89 (m, 4H), 2.80 (d, J=13.3Hz, 1H), 2.51 (s, 1H), 1.99 (t, J=12.7 Hz, 2H), 1.85 (d, J=11.6 Hz, 2H),1.75-1.43 (m, 2H), 1.25 (s, 2H).

Example 12 Synthesis of(9S)-2-(2-methylpyridin-4-yl)-N-(pyrimidin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.350 g, 1.314 mmol), DMAP (0.482 g, 3.94 mmol) in THF (7 mL) was addedphenyl pyrimidin-4-ylcarbamate (0.848 g, 3.94 mmol) at 25° C. Thereaction mixture was stirred at 80° C. for 16 h. The solvent was removedunder vacuo and crude was diluted with CH₂Cl₂ (20 ml) and washed withwater (5 mL), saturated brine solution (5 mL) and dried over Na₂SO₄. Theorganic solvent was evaporated under reduced pressure to obtain thecrude product. The crude mixture was purified by flash columnchromatography (silica-gel: 100-200 mesh, eluted with 3% MeOH in CH₂Cl₂)to afford(9S)-2-(2-methylpyridin-4-yl)-N-(pyrimidin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(207 mg, 0.529 mmol, 40.3% yield) as a white solid (TLC: 10% MeOH inDCM, R_(f): 0.7), LCMS (m/z): 388.3 [M+H]⁺.

¹H NMR (400 MHz, DMSO-d6): δ 14.32 (s, 1H), 8.95 (d, J=1.3 Hz, 1H), 8.69(d, J=5.8 Hz, 1H), 8.61 (d, J=5.3 Hz, 1H), 8.17 (d, J=1.7 Hz, 1H), 8.15(dd, J=5.8, 1.3 Hz, 1H), 7.95 (dd, J=5.3, 1.8 Hz, 1H), 7.86 (d, J=8.1Hz, 1H), 7.74 (d, J=8.1 Hz, 1H), 4.90-4.67 (m, 1H), 3.40 (dd, J=13.7,1.9 Hz, 1H), 3.28 (s, 2H), 3.00-2.81 (m, 1H), 2.64 (s, 3H), 2.13-1.71(m, 2H), 1.41-1.14 (m, 2H).

Example 13 Synthesis of(9S)-2-(2-methylpyridin-4-yl)-N-(pyrazin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4] diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.700 g, 2.63 mmol) in Tetrahydrofuran (THF) (30 mL) stirred undernitrogen at room temperature ° C. was added triethylamine (2.198 mL,15.77 mmol) and triphosgene (0.780 g, 2.63 mmol). Then reaction mixturewas stirred at room temperature for 30 minutes. pyrazin-2-amine (0.750g, 7.88 mmol) was added at rt. Then the reaction mixture was stirred at65° C. for 24 hr. The reaction mixture was concentrated to dryness andthen partitioned between water (20 mL) and Dichloromethane (50 mL).Organic layer was separated and was dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to give crude as brown solid. Thecrude was purified by column chromatography using (100-200 mesh) silicagel to afford pure(9S)-2-(2-methylpyridin-4-yl)-N-(pyrazin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamideas a pale yellow solid, LCMS (m/z): 388.3 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 14.27 (s, 1H), 9.59 (d, J=0.9 Hz, 1H),8.67-8.60 (m, 1H), 8.32 (d, J=1.0 Hz, 2H), 8.10 (dt, J=1.9, 0.7 Hz, 1H),7.74-7.67 (m, 1H), 7.62-7.50 (m, 2H), 5.06-5.00 (m, 1H), 3.51-3.27 (m,3H), 3.03 (d, J=13.6 Hz, 1H), 2.74 (s, 3H), 2.26 (s, 1H), 2.08-1.84 (m,1H), 1.43 (d, J=6.2 Hz, 2H).

Example 14 Synthesis of(9S)-2-(2-methylpyridin-4-yl)-N-(pyridazin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.150 g, 0.563 mmol), phenyl pyridazin-3-ylcarbamate (0.364 g, 1.690mmol) in THF (3 mL) was added DMAP (0.206 g, 1.690 mmol) at 25° C. Thereaction mixture was stirred at 80° C. for 16 h. Solvent was removedunder reduced pressure, crude compound was diluted with DCM (10 ml),washed with water (3 ml), saturated brine solution (3 ml), dried overNa₂SO₄ and concentrated under reduced pressure to obtain crude product.The crude mixture was purified by flash column chromatography(silica-gel: 100-200 mesh, eluent: 2% MeOH in CH₂Cl₂) to afford(9S)-2-(2-methylpyridin-4-yl)-N-(pyridazin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(271 mg, 0.692 mmol, 123% yield) as a white solid (TLC: 10% MeOH inCH₂Cl₂, R_(f): 0.6), LCMS (m/z): 388.3 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 14.66 (s, 1H), 8.94 (dd, J=4.7, 1.5 Hz, 1H),8.66 (dd, J=5.2, 0.8 Hz, 1H), 8.50 (dd, J=9.1, 1.5 Hz, 1H), 8.29 (d,J=1.8 Hz, 1H), 7.74 (dd, J=5.4, 1.8 Hz, 1H), 7.57 (d, J=4.3 Hz, 2H),7.46 (dd, J=9.0, 4.7 Hz, 1H), 5.00 (p, J=2.6 Hz, 1H), 3.50-3.22 (m, 3H),3.12-2.96 (m, 1H), 2.82 (s, 3H), 2.39-2.10 (m, 1H), 1.95 (tdd, J=14.2,5.5, 3.0 Hz, 1H), 1.58-1.33 (m, 2H).

Example 15 Synthesis of(9S)-N-(2-methyl-2H-indazol-5-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

DMAP (0.550 g, 4.51 mmol) was added to a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.4 g, 1.502 mmol) and phenyl (2-methyl-2H-indazol-5-yl)carbamate(0.803 g, 3.00 mmol) in Tetrahydrofuran (THF) (30 mL) under nitrogenatmosphere at room temp. The reaction was stirred at 70° C. for 48 hr.Reaction was cooled to room temperature and concentrated under reducedpressure. Added EtOAc to reaction mass and stirred for 10 minutes beforebeing filtered and the solids washed with EtOAc. Take filtrate andconcentrated to give the crude as brown solid (TLC eluent: 80% EtOAc:R_(f)-0.4; UV active). The crude was purified by column chromatographyusing neutral alumina and was eluted with 60% EtOAc in Hexane to affordpure afford(9S)-N-(2-methyl-2H-indazol-5-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.18 g, 0.406 mmol, 27.1% yield) as a off-white solid, LCMS (m/z):440.4 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.44 (s, 1H), 8.66 (d, J=5.2 Hz, 1H), 8.18(dd, J=1.9, 0.8 Hz, 1H), 7.82 (d, J=0.9 Hz, 1H), 7.69 (d, J=1.6 Hz, 1H),7.63 (d, J=9.1 Hz, 1H), 7.60-7.50 (m, 2H), 7.40 (d, J=7.9 Hz, 1H),7.27-7.19 (m, 1H), 5.05 (t, J=2.7 Hz, 1H), 4.20 (s, 3H), 3.50-3.23 (m,3H), 3.01 (d, J=13.5 Hz, 1H), 2.67 (s, 3H), 2.28 (s, 1H), 1.93 (s, 1H),1.58 (s, 2H).

Example 16 Synthesis of(9S)-2-(2-methylpyridin-4-yl)-N-(pyrimidin-5-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.6 g, 2.253 mmol), triethylamine (0.942 mL, 6.76 mmol) andbis(trichloromethyl) carbonate (0.334 g, 1.126 mmol) in THF (12 mL)stirred under nitrogen at room temperature for 30 min. min and thenadded pyrimidin-5-amine (0.643 g, 6.76 mmol) in one charge. The reactionmixture was stirred at 65° C. for 16 h. The solvent was removed underreduced pressure and the obtained crude was diluted with CH₂Cl₂ (35 ml),washed with water (5 mL), saturated brine solution (5 mL), dried overanhydrous Na₂SO₄ and concentrated under reduced pressure to obtain thecrude product. The crude mixture was purified by flash columnchromatography (silica-gel: 100-200 mesh, eluent: 3% MeOH in CH₂Cl₂) toobtained semi pure compound. The semi pure compound was again purifiedby Prep HPLC (30% ACN/Water) to afford(9S)-2-(2-methylpyridin-4-yl)-N-(pyrimidin-5-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(261 mg, 0.667 mmol, 29.6% yield) as an white solid (TLC: 10% MeOH inCH₂Cl₂, R_(f): 0.4), LCMS (m/z): 388 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.81 (s, 1H), 9.03 (s, 2H), 8.94 (s, 1H),8.69 (dd, J=5.2, 0.9 Hz, 1H), 7.75-7.55 (m, 2H), 7.55-7.46 (m, 1H), 7.43(d, J=8.0 Hz, 1H), 5.08-4.89 (m, 1H), 3.49-3.17 (m, 3H), 3.02 (d, J=13.7Hz, 1H), 2.69 (s, 3H), 2.36-2.13 (m, 1H), 2.06-1.79 (m, 1H), 1.59-1.32(m, 2H).

Example 17 Synthesis of(9S)-N-(pyrazin-2-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-2-(2-(trifluoromethyl)pyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.9 g, 2.81 mmol) in THF (18 mL) was added bis(trichloromethyl)carbonate (0.417 g, 1.405 mmol) at RT and stirred for 30 min. ThenPyrazine-2-amine and Et₃N (2.94 ml, 21.8 mmol) were added. The reactionmixture was stirred at 65° C. for 16 h. The organic solvent was removedunder reduced pressure and CH₂Cl₂ (30 ml). The organic layer was washedwith water (5 ml), saturated brine solution (5 ml), dried over anhydrousNa₂SO₄ and concentrated under reduced pressure to obtain the crudeproduct. The crude mixture was purified by flash column chromatography(silica-gel: 100-200 mesh, eluent: 4% MeOH in DCM) to afford(9S)-N-(pyrazin-2-yl)-2-(2-(trifluoromethyl)pyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(476 mg, 1.059 mmol, 37.7% yield) as a white solid (TLC: 80% EtOAc inHexane, R_(f): 0.5), LCMS (m/z): 442.2 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 14.06 (s, 1H), 9.56 (d, J=1.4 Hz, 1H), 8.87(d, J=5.1 Hz, 1H), 8.46 (dd, J=1.8, 0.9 Hz, 1H), 8.37-8.23 (m, 2H), 8.14(dd, J=5.1, 1.7 Hz, 1H), 7.63 (d, J=8.0 Hz, 1H), 7.56 (s, 1H), 5.15-4.92(m, 1H), 3.52-3.26 (m, 3H), 3.03 (d, J=14.0 Hz, 1H), 2.26 (s, 1H),2.05-1.88 (m, 1H), 1.53-1.29 (m, 2H).

Example 18 Synthesis of(9S)-N-(pyrimidin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak1 from the intermediate SFC separation 500 mg, 1.532 mmol) intetrahydrofuran (500 mL), DMAP (562 mg, 4.60 mmol) and phenyl2-(pyrimidin-2-yl)acetate (985 mg, 4.60 mmol) were added. The reactionmixture was stirred at 65° C. for 28 hr. The reaction mixture wasdiluted with water (50 mL) and extracted with EtOAc (2×75 mL). Thecombined organic layer was washed with water and saturated with brinesolution and dried over anhydrous Na₂SO₄, filtered and concentrated togive the crude as white solid (TLC eluent: 10% MeOH in DCM: R_(f)-0.4;UV active). The crude product was purified by column chromatography(neutral alumina) and the product was eluted with 25% ethyl acetate inhexane to afford(9S)-N-(pyrimidin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methano-pyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(320 mg, 0.711 mmol, 46.4% yield) as off white solid. LCMS (m/z) 448.3[M+H]⁺.

¹H NMR (400 MHz, DMSO-d₆): δ 13.80 (s, 1H), 8.62 (d, J=4.8 Hz, 2H), 7.36(d, J=8.6 Hz, 1H), 7.15 (t, J=4.8 Hz, 1H), 6.61 (d, J=8.6 Hz, 1H), 4.69(s, 1H), 4.30 (d, J=12.7 Hz, 1H), 4.20 (d, J=13.0 Hz, 1H), 3.26 (d,J=1.9 Hz, 1H), 3.20-2.89 (m, 4H), 2.79 (d, J=13.4 Hz, 1H), 2.55 (d,J=10.8 Hz, 1H), 1.99 (d, J=12.6 Hz, 2H), 1.82 (h, J=6.1 Hz, 2H),1.72-1.44 (m, 2H), 1.42-1.15 (m, 2H).

Example 19 Synthesis of(9S)-N-(4,5-dimethylthiazol-2-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(600 mg, 2.253 mmol) in THF (15 mL) were added triethylamine (0.942 mL,6.76 mmol) and triphosgene (334 mg, 1.126 mmol) at 30° C. and stirredfor 1 h. Then 4,5-dimethylthiazol-2-amine hydrochloride (556 mg, 3.38mmol) was added at 30° C. and reaction was heated at 70° C. for 16 h.The solvent evaporated under reduced pressure, residue diluted withwater (40 ml) and extracted with DCM (2×40 ml). The combined organiclayer was washed with water, brine, dried over anhydrous sodium sulfateand the solvent was evaporated under reduced pressure to obtain crudecompound. The crude mixture was purified by flash column chromatographyand prep HPLC to afford(9S)-N-(4,5-dimethylthiazol-2-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(275 mg, 0.655 mmol, 42% yield) as a pale yellow solid (TLC: 10% MeOH inEtOAc, R_(f): 0.3), LCMS (m/z): 421.27 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ ppm 14.79 (s, 2H), 8.64 (d, J=5.26 Hz, 2H),8.02 (s, 1H), 7.64 (dd, J=5.26, 1.53 Hz, 1H), 7.54 (q, J=8.11 Hz, 1H),4.99 (s, 1H), 3.42-3.18 (m, 3H), 3.12-2.89 (m, 1H), 2.79 (s, 3H), 2.28(s, 3H), 2.22 (s, 3H), 2.00-1.75 (m, 1H), 1.52-1.35 (m, 2H).

Example 20 Synthesis of(9S)-N-(6-methoxypyrazin-2-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.6 g, 2.253 mmol) in Tetrahydrofuran (THF) (30 mL) stirred undernitrogen at room temperature was added triethylamine (1.884 mL, 13.52mmol) and triphosgene (0.669 g, 2.253 mmol). Then reaction mixture wasstirred at room temperature for 30 minutes. 6-methoxypyrazin-2-amine(0.846 g, 6.76 mmol) was added at rt. Then the reaction mixture wasstirred at 65° C. for 16 hr. The reaction mixture was cooled to roomtemperature and concentrated under reduced pressure and was partitionedbetween water (20 mL) and EtOAc (50 mL). Organic layer was separated andwas dried over anhydrous Na₂SO₄, filtered and filtrate was evaporated togive crude as brown solid (TLC eluent: 100% EtOAc: R_(f)-0.3; UVactive). The crude was purified by column chromatography using neutralalumina and was eluted with 50% EtOAc in Hexane to afford pure(9S)-N-(6-methoxypyrazin-2-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.220 g, 0.526 mmol, 23.34% yield) as a off-white solid, LCMS (m/z):418.3 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.64 (s, 1H), 9.45-8.78 (m, 1H), 8.60 (dd,J=5.3, 0.8 Hz, 1H), 7.96 (d, J=0.5 Hz, 1H), 7.80 (ddd, J=5.3, 1.7, 0.7Hz, 1H), 7.62 (dt, J=1.8, 0.7 Hz, 1H), 7.59 (s, 1H, 7.57 (s, 1H), 7.46(d, J=8.0 Hz, 1H), 5.04 (t, J=2.6 Hz, 1H), 3.84 (s, 3H), 3.57-3.26 (m,3H), 3.15-2.91 (m, 1H), 2.64 (s, 3H), 2.28 (d, J=14.3 Hz, 1H), 2.12-1.85(m, 1H), 1.43 (s, 2H)

Example 21 Synthesis of((9S)-N-(6-methylpyrazin-2-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Triphosgene (0.279 g, 0.939 mmol) was added to a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.25 g, 0.939 mmol) in tetrahydrofuran (50 mL) at 25° C. Triethylamine(0.654 mL, 4.69 mmol) was added and followed by addition of6-methylpyrazin-2-amine (0.410 g, 3.75 mmol). The reaction mixture wasstirred for 15 h at 70° C. The reaction mixture was cooled to 28° C. andwas partitioned between water (50 mL) and dichloromethane (50 mL). Theseparated organic layer was washed with water and brine. The organiclayer was dried over sodium sulfate filtered and filtrate was evaporatedto get crude compound (TLC eluent: 10% MeOH in EtOAc; R_(f)=0.4; UVactive). The crude compound was purified by using neutral alumina andeluted in 100% ethyl acetate to afford(9S)-N-(6-methylpyrazin-2-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (0.11 g, 0.272 mmol, 29.0% yield) asyellow solid, LCMS (m/z): 402.26 [M+H]⁺.

¹H NMR (CDCl₃, 400 MHz): δ 14.02 (s, 1H), 9.37 (s, 1H), 8.68 (d, J=5.26Hz, 1H), 8.13-8.26 (m, 1H), 7.98 (dd, J=5.26, 1.75 Hz, 1H), 7.69-7.80(m, 1H), 7.49-7.63 (m, 2H), 5.03 (t, J=2.19 Hz, 1H), 3.30-3.47 (m, 2H),3.03 (br d, J=13.81 Hz, 1H), 2.69 (s, 2H), 2.49-2.59 (m, 2H), 2.15-2.30(m, 1H), 1.84-1.99 (m, 1H), 1.24-1.66 (m, 3H).

Example 22 Synthesis of(9S)-N-(pyrimidin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak II from the intermediate SFC separation 0.600 g, 1.838 mmol) inTetrahydrofuran (THF) (30 mL) was added DMAP (0.674 g, 5.52 mmol) andphenyl pyrimidin-2-ylcarbamate (1.187 g, 5.52 mmol) stirred at 65° C.for 36 hr. Reaction was cooled to room temperature and then concentratedunder reduced pressure. The residue was partitioned between water (10mL) and EtOAc (50 mL). Organic layer was separated and was dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude asbrown solid (TLC eluent: 80% EtOAc in Hexane: R_(f)-0.3; UV active). Thecrude was purified by column chromatography using neutral alumina andwas eluted with 35-40% EtOAc in Hexane to afford pure9S)-N-(pyrimidin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.375 g, 0.834 mmol, 45.4% yield) as a off-white solid, LCMS (m/z):448.3 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.97 (s, 1H), 8.59 (d, J=4.8 Hz, 2H), 7.31(d, J=8.6 Hz, 1H), 6.93 (t, J=4.8 Hz, 1H), 6.40 (d, J=8.6 Hz, 1H), 4.93(q, J=2.6 Hz, 1H), 4.44-4.07 (m, 2H), 3.31 (d, J=1.9 Hz, 1H), 3.19 (d,J=3.4 Hz, 2H), 3.10-2.81 (m, 3H), 2.44 (dtt, J=15.6, 7.7, 3.9 Hz, 1H),2.29 (d, J=13.4 Hz, 1H), 2.17-2.07 (m, 1H), 1.93 (dt, J=13.2, 3.3 Hz,1H), 1.85 (ddd, J=13.8, 5.3, 3.1 Hz, 2H), 1.77-1.68 (m, 1H), 1.55-1.43(m, 1H), 1.31 (d, J=14.0 Hz, 1H).

Example 23 Synthesis of(9S)-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

NaH (0.154 g, 6.43 mmol) was added to a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]d

iazocine (Peak-I from intermediate SFC Separation, 0.350 g, 1.072 mmol)in tetrahydrofuran (25 mL) stirred under nitrogen at 0° C. The reactionmixture was stirred at 30° C. for 30 minutes and3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (0.386 g,1.609 mmol) was added. The reaction mixture was stirred at 65° C. for 24hrs. The reaction mixture was cooled to room temperature and quenchedwith methanol (5 mL). The reaction mixture was diluted with water andextracted with EtOAc (2×100 mL). The organic layers were washed withwater and followed by brine solution and dried over with Na₂SO₄ filteredand concentrated under reduced pressure to get crude as a brown solid.The crude was purified by column chromatography using Neutral Aluminaand was eluted with 60% ethyl acetate in hexane to afford pure(9S)-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.320 g, 0.708 mmol, 66.0% yield) as off white solid, LCMS (m/z):447.34 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.58 (s, 1H), 8.46-8.04 (m, 2H), 7.65 (ddd,J=8.9, 7.3, 1.9 Hz, 1H), 7.35-7.10 (m, 1H), 6.94 (ddd, J=7.3, 4.9, 1.1Hz, 1H), 6.38 (d, J=8.6 Hz, 1H), 5.10-4.73 (m, 1H), 4.59-4.37 (m, 1H),4.31-4.19 (m, 1H), 3.30 (dd, J=13.4, 1.9 Hz, 1H), 3.26-3.13 (m, 2H),3.06-2.88 (m, 3H), 2.48-2.36 (m, J=16.3, 8.3, 4.1 Hz, 1H), 2.21 (d,J=14.4 Hz, 1H), 2.12 (d, J=12.1 Hz, 1H), 1.94 (s, 1H), 1.88-1.68 (m,2H), 1.67-1.57 (m, 1H), 1.50 (s, 1H), 1.31 (d, J=13.6 Hz, 1H).

Example 24 Synthesis of(9S)-N-(5-fluoropyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-II of the intermediate SFC separation, 0.600 g, 1.838 mmol) inTetrahydrofuran (THF) (30 mL) was added DMAP (0.674 g, 5.52 mmol) andphenyl (5-fluoropyridin-2-yl)carbamate (1.281 g, 5.52 mmol). Thereaction mixture was stirred at 65° C. for 36 hr. Reaction was cooled toroom temperature and concentrated under reduced pressure. The residuewas partitioned between water (10 mL) and EtOAc (50 mL). Organic layerwas separated and was dried over anhydrous Na₂SO₄, filtered and filtratewas evaporated to give crude as brown solid (TLC eluent: 80% EtOAc inHexane: R_(f)-0.3; UV active). The crude was purified by columnchromatography using neutral alumina and was eluted with 25-30% EtOAc inHexane to afford pure(9S)-N-(5-fluoropyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.288 g, 0.608 mmol, 33.1% yield) as a white solid, LCMS (m/z): 465.3[M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.62 (s, 1H), 8.20 (ddd, J=9.2, 4.1, 0.6 Hz,1H), 8.09 (s, 1H), 7.39 (ddd, J=9.1, 7.8, 3.0 Hz, 1H), 7.30 (d, J=8.5Hz, 1H), 6.39 (d, J=8.6 Hz, 1H), 4.87 (p, J=2.8 Hz, 1H), 4.30 (dtt,J=12.5, 3.7, 1.8 Hz, 2H), 3.31 (dd, J=13.4, 1.9 Hz, 1H), 3.25-3.13 (m,2H), 3.07-2.85 (m, 3H), 2.44 (dt, J=7.4, 3.6 Hz, 1H), 2.26-2.05 (m, 2H),1.99-1.83 (m, 2H), 1.78-1.66 (m, 1H), 1.62 (dd, J=12.4, 3.8 Hz, 1H),1.53-1.44 (m, 1H), 1.37-1.26 (m, 1H).

Example 25 Synthesis of(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-I of the intermediate SFC separation, 600 mg, 1.838 mmol) intetrahydrofuran (30 mL), DMAP (674 mg, 5.52 mmol) and phenylpyridin-3-ylcarbamate (1182 mg, 5.52 mmol) were added. The reactionmixture was stirred at 65° C. for 16 hr. The reaction mixture wasdiluted with water (50 mL) and extracted with ethyl acetate (2×75 mL).The combined organic layers were washed with water and saturated withbrine solution and dried over anhydrous sodium sulfate, filtered andconcentrated to give crude as a white solid. (TLC eluent: 100% pureEthyl Acetate; R_(f) value: 0.4; UV active). The crude product waspurified by column chromatography (neutral alumina) product was elutedwith 25% ethyl acetate in hexane to afford(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(350 mg, 0.782 mmol, 42.5% yield) as off white solid, LCMS (m/z): 447.26[M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 12.94 (s, 1H), 8.62 (dd, J=2.6, 0.7 Hz, 1H),8.32 (dd, J=4.7, 1.5 Hz, 1H), 8.15 (ddd, J=8.4, 2.7, 1.5 Hz, 1H), 7.33(d, J=8.5 Hz, 1H), 7.29-7.24 (m, 1H), 6.41 (d, J=8.6 Hz, 1H), 4.90 (t,J=2.6 Hz, 1H), 4.22-4.04 (m, 2H), 3.31 (dd, J=13.5, 1.9 Hz, 1H),3.25-3.14 (m, 2H), 2.99-2.88 (m, 3H), 2.48-2.36 (m, 1H), 2.25-2.18 (m,1H), 2.13 (dd, J=13.2, 4.0 Hz, 1H), 1.98-1.92 (m, 1H), 1.85 (tdd,J=13.9, 5.2, 2.9 Hz, 1H), 1.71 (dtd, J=16.5, 8.3, 7.7, 4.3 Hz, 1H), 1.63(d, J=3.8 Hz, 1H), 1.57-1.44 (m, 1H), 1.33 (d, J=14.2 Hz, 1H).

Example 26 Synthesis of(9S)-N-(1-methyl-1H-pyrazol-4-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

DMAP (0.573 g, 4.69 mmol) was added to a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(0.5 g, 1.877 mmol), and phenyl (1-methyl-1H-pyrazol-4-yl)carbamate(1.01 g, 4.692 mmol) in tetrahydrofuran (10 mL) at room temperature andheated to 24 h at 80° C. in sealed tube. The reaction mixture was cooledto 28° C., and was diluted with ethyl acetate and water. The organiclayer was separated and was washed with water and brine. The organiclayer was dried over sodium sulfate and evaporated under reducedpressure to get crude compound (TLC eluent: 10% MeOH in EtOAc;R_(f)-0.4; UV active). The crude compound was purified by using neutralalumina and was eluted with 75% ethyl acetate in hexane to afford(9S)-N-(1-methyl-1H-pyrazol-4-yl)-2-(2-methylpyridin-4-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]dia-zocine-10(7H)-carboxamide(0.17 g, 0.430 mmol, 22.89% yield) as an off-white solid, LCMS (m/z):390.28 [M+H]⁺.

¹H NMR (CDCl₃, 400 MHz): δ 13.27 (s, 1H), 8.67 (d, J=5.26 Hz, 1H), 7.87(s, 1H), 7.63 (d, J=1.53 Hz, 1H), 7.53-7.57 (m, 2H), 7.43 (d, J=0.66 Hz,1H), 7.40 (s, 1H), 4.99 (t, J=2.30 Hz, 1H), 3.89 (s, 3H), 3.22-3.42 (m,3H), 2.98 (br d, J=13.59 Hz, 1H), 2.70 (s, 3H), 2.20-2.28 (m, 1H), 1.92(tdd, J=13.67, 13.67, 5.54, 3.07 Hz, 1H), 1.32-1.50 (m, 2H).

Example 27 Synthesis of(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

Diphenyl phosphorazidate (1.996 g, 7.25 mmol) added to a stirredsolution of pyrazine-2-carboxylic acid (Peak-II of the intermediate SFCseparation, 0.600 g, 4.83 mmol) and DIPEA (4.22 mL, 24.17 mmol) inTetrahydrofuran (THF) (60 mL) stirred with argon at room temp. Thereaction mixture was stirred 2 hr at room temperature and added(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(1.105 g, 3.38 mmol). The reaction mixture was stirred 16 hr at 65° C.The reaction mixture was cooled to room temperature and concentratedunder reduced pressure. The residue was partitioned between water (30mL) and EtOAc (100 mL). Organic layer was separated and was dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude asbrown solid (TLC eluent: 100% EtOAc: R_(f)-0.3; UV active). The crudewas purified by column chromatography using neutral alumina and waseluted with 50% EtOAc in Hexane to afford pure(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.367 g, 0.815 mmol, 16.86% yield) as a pale yellow solid, LCMS (m/z):448.3 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 13.79 (s, 1H), 9.53 (d, J=1.5 Hz, 1H), 8.25(d, J=2.5 Hz, 1H), 8.20 (d, J=1.6 Hz, 1H), 7.32 (d, J=8.6 Hz, 1H), 6.42(d, J=8.6 Hz, 1H), 4.90 (s, 1H), 4.28 (s, 2H), 3.32 (dd, J=13.5, 1.9 Hz,1H), 3.21 (dd, J=12.5, 3.3 Hz, 2H), 3.10-2.86 (m, 3H), 2.44 (dt, J=7.6,3.6 Hz, 1H), 2.28-2.18 (m, 1H), 2.11 (d, J=3.6 Hz, 1H), 1.99-1.80 (m,2H), 1.78-1.65 (m, 1H), 1.65-1.60 (m, 1H), 1.49 (s, 1H), 1.35 (s, 1H).

Example 28 Synthesis of(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-II of the intermediate SFC separation, (Peak-II from intermediateSFC separation, 0.700 g, 2.145 mmol) in Tetrahydrofuran (THF) (70 mL)was added DMAP (0.786 g, 6.43 mmol) and phenyl pyridin-3-ylcarbamate(1.378 g, 6.43 mmol) stirred at 65° C. for 36 hr before the reaction wascooled to room temperature and concentrated under reduced pressure. Theresidue was partitioned between water (20 mL) and EtOAc (75 mL). Organiclayer was separated and was dried over anhydrous Na₂SO₄, filtered andfiltrate was evaporated to give crude as brown solid (TLC eluent: 80%EtOAc in Hexane: R_(f)-0.3; UV active). The crude was purified by columnchromatography using neutral alumina and was eluted with 50% EtOAc inHexane to afford pure(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.355 g, 0.794 mmol, 37.0% yield) as a white solid, LCMS (m/z): 447.3[M+H]⁺.

¹H-NMR (400 MHz, CDCl₃): δ ppm 12.92 (s, 1H), 8.62 (d, J=2.63 Hz, 1H),8.32 (dd, J=4.71, 1.43 Hz, 1H), 8.08-8.21 (m, 1H), 7.33 (d, J=8.55 Hz,1H), 7.20-7.29 (m, 1H), 6.40 (d, J=8.55 Hz, 1H), 4.90 (br s, 1H),4.00-4.21 (m, 2H), 3.31 (dd, J=13.48, 1.86 Hz, 1H), 3.17-3.26 (m, 2H),2.88-3.03 (m, 3H), 2.36-2.49 (m, 1H), 2.19-2.25 (m, 1H), 2.10-2.16 (m,1H), 1.92-1.99 (m, 1H), 1.86 (tdd, J=13.84, 13.84, 5.10, 2.96 Hz, 1H),1.65-1.76 (m, 1H), 1.46-1.58 (m, 2H), 1.29-1.38 (m, 1H)

Example 29 Synthesis of(9S)-N-(5-fluoropyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-I from SFC separation, 450 mg, 1.379 mmol) in tetrahydrofuran (30mL), DMAP (505 mg, 4.14 mmol) and phenyl (5-fluoropyridin-2-yl)carbamate(961 mg, 4.14 mmol) were added. The reaction mixture was stirred at 65°C. for 16 hr. The reaction mixture was diluted with water (50 mL) andextracted with ethyl acetate (2×75 mL). The combined organic layers werewashed with water and saturated with brine solution and dried overanhydrous sodium sulfate, filtered and concentrated to give the crude asoff white solid. (TLC Eluent: 100% Ethyl Acetate; R_(f) value: 0.4; UVactive). The crude product was purified by column chromatography(neutral alumina) product was eluted with 25% ethyl acetate in hexane toafford(9S)-N-(5-fluoropyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(140 mg, 0.296 mmol, 21.46% yield) as a white solid, LCMS (m/z): 465.21[M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ ppm 1.15-1.43 (m, 1H) 1.43-1.68 (m, 2H)1.68-1.89 (m, 3H) 2.04-2.28 (m, 2H) 2.42 (dtd, J=11.67, 7.92, 7.92, 4.06Hz, 1H) 2.86-3.09 (m, 3H) 3.10-3.36 (m, 3H) 4.23-4.47 (m, 2H) 4.87 (brs, 1H) 6.39 (d, J=8.55 Hz, 1H) 7.3 (d, J=8.8 Hz, 1H) 7.34-7.59 (m, 1H)8.09 (d, J=2.8 Hz, 1H) 8.20 (dd, J=9.21, 4.17 Hz, 1H) 13.65 (s, 1H).

Example 30 Synthesis of(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

A suspension of pyrazine-2-carboxylic acid (0.6 g, 4.83 mmol) intetrahydrofuran (50 mL) diphenylphosphoryl azide (1.996 g, 7.25 mmol)and DIPEA (4.22 mL, 24.17 mmol) added to the reaction mixture. Thereaction mixture was stirred for 2 hr at 28° C.(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diaz-ocine(Peak-I from intermediate SFC Separation 1.105 g, 3.38 mmol) was addedto the reaction mixture. The reaction mixture was stirred 16 hr at 65°C. The reaction mixture was diluted with water (50 mL) and extractedwith ethyl acetate (2×75 mL). The combined organic layer was washed withwater and saturated with brine solution and dried over anhydrous Na₂SO₄,filtered and concentrated to give the crude as a off white solid. (TLCeluent: 100% ethyl acetate; R_(f) value: 0.3; UV active). The crudeproduct was purified by column chromatography (neutral alumina) productwas eluted with 30% ethyl acetate in hexane to afford(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)-piperidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.330 g, 0.728 mmol, 15.05% yield) as a white solid, LCMS (m/z): 448.25[M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ ppm 1.31-1.34 (m, 1H) 1.45-1.67 (m, 2H)1.68-1.79 (m, 1H) 1.82-1.99 (m, 2H) 2.05-2.18 (m, 2H) 2.42 (dddt,J=15.65, 11.78, 8.03, 3.97, 3.97 Hz, 1H) 2.86-3.09 (m, 3H) 3.11-3.35 (m,3H) 4.21-4.39 (m, 2H) 4.89 (br s, 1H) 6.42 (d, J=8.55 Hz, 1H) 7.32 (d,J=8.4 Hz, 1H) 8.14-8.20 (m, 1H) 8.24 (d, J=2.8 Hz, 1H) 9.53 (d, J=1.53Hz, 1H) 13.81 (s, 1H)

Example 31 Synthesis of(9S)-N-(pyridine-2-yl)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamidehydrochloride

2.0 M Hydrochloric acid in Diethyl ether (2 mL, 4.00 mmol) was added to(9S)-N-(Pyridine-2-yl)-2-(3-(trifluoromethyl)pyrrolidine-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(180 mg, 0.416 mmol) at 0° C. The reaction mixture was stirred at 28° C.for 4 h and concentrated to give crude compound (TLC eluent: 5% MeOH inDCM: R_(f)-0.1; UV active). The crude compound was washed with Diethylether (2×5 mL) to afford pure(9S)-N-(pyridine-2-yl)-2-(3-(trifluoromethyl)pyrrolidin-1-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamidehydrochloride (83 mg, 0.177 mmole, 42.78% yield) as pale brown solid,LCMS (m/z): 433.28 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 15.02 (br s, 1H), 13.34 (br s, 1H), 8.25 (brd, J=3.73 Hz, 1H), 8.13 (br d, J=8.33 Hz, 1H), 8.06 (d, J=8.77 Hz, 1H),7.75-7.68 (m, 1H), 7.04-6.98 (m, 1H), 6.15 (s, 1H), 5.30-5.24 (b, s,1H), 4.12-3.94 (m, 2H), 3.73-3.59 (m, 4H), 3.49-3.37 (m, 2H), 3.18-3.04(m, 1H), 2.45-2.25 (m, 3H), 1.86 (d, J=12.50 Hz, 2H).

Example 32 Synthesis of(9S)-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(300 mg, 1.126 mmol) in THF (30 mL) under nitrogen atmosphere at RT wasadded phenyl pyridin-3-ylcarbamate (724 mg, 3.38 mmol), DMAP (413 mg,3.38 mmol) and stirred at 65° C. for 48 h. (TLC eluent: 5% MeOH in DCM:R_(f)-0.4; UV active). The reaction mixture was cooled to RT,concentrated and the residue was partitioned between water (20 mL) andEtOAc (100 mL). Organic layer was separated, dried over anhydrousNa₂SO₄, filtered and filtrate was evaporated to give crude compound. Thecrude was purified by column chromatography (neutral alumina, eluent: 2%MeOH in DCM) to afford the desired product(9S)-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.516 mmol, 45.8% yield) as a pale yellow solid. LCMS (m/z):387.09 [M+H]⁺, R_(t)=1.21 min.

¹H NMR (400 MHz, CDCl₃): ppm 13.61 (s, 1H), 8.84-8.62 (m, 2H), 8.33 (dd,J=4.71, 1.21 Hz, 1H), 8.17 (dd, J=8.33, 1.54 Hz, 1H), 7.63 (s, 1H), 7.58(d, J=7.89 Hz, 1H), 7.52 (br d, J=5.04 Hz, 1H), 7.42 (d, J=8.11 Hz, 1H),7.28 (br d, J=4.60 Hz, 1H), 5.02 (br s, 1H), 3.45-3.31 (m, 3H), 3.01 (brd, J=13.59 Hz, 1H), 2.69 (s, 3H), 2.27 (br d, J=12.93 Hz, 1H), 3.01-1.85(m, 1H), 1.54-1.32 (m, 2H).

Example 33 Synthesis of(9S)-3-chloro-N2-cyclopropyl-N10-(pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (650 mg, 1.739 mmol) in DMF (6 mL), under nitrogen at RT was addedDIPEA (0.911 mL, 5.22 mmol), HATU (1322 mg, 3.48 mmol) andcyclopropanamine (119 mg, 2.087 mmol), then the reaction mixture wasstirred for 16 h. (TLC system: 5% Methanol in DCM. R_(f) value: 0.30).Reaction mass was diluted with 50 mL of ice cold water, extracted withEtOAc (3×100 mL). The combined organic layer was washed with brine (100mL), dried over Na₂SO₄, filtered and concentrated to get crude compound.The crude material was purified by combiflash (using silica gel column,75% EtOAc in Hexane). Fractions containing pure compound were combinedand concentrated to afford the desired compound to get(9S)-3-chloro-N2-cyclopropyl-N10-(pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-H)-dicarboxamide(97 mg, 0.229 mmol, 13.15% yield) as off white solid. LCMS (m/z): 413.10[M+H]⁺, Rt=1.84 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.12 (br s, 1H), 8.33 (dd, J=4.93, 0.99Hz, 1H), 8.14 (d, J=8.33 Hz, 1H), 7.51-7.76 (m, 2H), 7.60 (m, 1H), 7.09(br, 1H), 4.97 (br s, 1H), 3.23-3.44 (m, 3H), 3.04 (ddt, J=10.91, 7.18,3.89, 3.89 Hz, 1H), 2.93 (br d, J=13.81 Hz, 1H), 2.10-2.32 (m, 1H),1.80-2.07 (m, 1H), 1.22-1.46 (m, 2H), 0.81-0.98 (m, 4H)

Example 34 Synthesis of(9S)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(200 mg, 0.354 mmol) in Methanol (10 mL) was added aq.HCl (0.308 mL,3.54 mmol), drop wise over a period of 5 min at 0° C. and stirred atroom temperature for 1 h. (TLC system: 100% Ethyl acetae. Rf value:0.3). Then the reaction mixture was quenched with saturated NaHCO₃solution (20 mL) and extracted with DCM (2×30 mL). The combined organiclayer was dried over anhydrous sodium sulphate and concentrated underreduced pressure to obtain crude product. The crude product wastriturated with pentane: diethyl ether (1:1) to afford the requiredpurity of desired product(9S)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(100 mg, 0.187 mmol, 52.7% yield) as an off white solid. LCMS (m/z):525.14 [M+H]⁺, Rt=1.53 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.84 (s, 1H), 8.16 (br d, J=9.21 Hz,1H), 8.09 (d, J=5.92 Hz, 1H), 8.00 (d, J=7.89 Hz, 1H), 7.80 (d, J=2.19Hz, 1H), 7.61 (d, J=7.89 Hz, 1H), 6.60 (dd, J=5.81, 2.30 Hz, 1H),5.17-5.04 (m, 1H), 4.96 (br s, 1H), 4.25-4.03 (m, 3H), 3.89-3.71 (m,2H), 3.44-3.18 (m, 3H), 2.96 (br d, J=14.03 Hz, 1H), 2.59 (br s, 1H),2.22 (br d, J=14.47 Hz, 1H), 2.06-1.76 (m, 2H), 1.60 (d, J=7.02 Hz, 3H),1.45-1.33 (m, 2H).

Example 35 Synthesis of(9S)-N2-(2,2-difluoropropyl)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-N2-(2,2-difluoropropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(210 mg, 0.384 mmol) in Methanol (10 mL) was added aq.HCl (0.667 mL,7.68 mmol) drop wise over a period of 5 min at 0° C. and stirred at roomtemperature for 1 h. (TLC system: 100% Ethyl acetae. Rf value: 0.3).Then, the reaction mixture was quenched with saturated NaHCO₃ solution(20 mL) and extracted with DCM (2×30 mL). The combined organic layer wasdried over anhydrous sodium sulphate and concentrated under reducedpressure and was triturated with pentane: diethyl ether (1:1) to affordthe desired product(9S)-N2-(2,2-difluoropropyl)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(95 mg, 0.186 mmol, 48.5% yield) as an off white solid. LCMS (m/z):507.14 [M+H]⁺, Rt=1.40 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 14.25 (s, 1H), 8.69 (br t, J=6.47 Hz,1H), 8.12 (d, J=5.92 Hz, 1H), 7.95 (d, J=8.11 Hz, 1H), 7.71 (d, J=2.19Hz, 1H), 7.60 (d, J=7.89 Hz, 1H), 6.58 (dd, J=5.92, 2.41 Hz, 1H), 4.94(br s, 1H), 4.24-4.08 (m, 3H), 4.04-3.90 (m, 2H), 3.87-3.73 (m, 2H),3.43-3.24 (m, 3H), 2.97 (br d, J=14.03 Hz, 1H), 2.62-2.50 (m, 1H),2.27-2.16 (m, 1H), 2.08-1.89 (m, 2H), 1.69 (t, J=18.64 Hz, 3H),1.465-1.35 (m, 2H).

Example 36 Synthesis of(9S)-N2-cyclopropyl-N10-(pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid(6 g, 17.68 mmol) in DMF (60 mL), under nitrogen at 0° C. was addedDIPEA (9.26 mL, 53.0 mmol), HATU (13.45 g, 35.4 mmol) andcyclopropanamine (1.514 g, 26.5 mmol) and the reaction mixture wasstirred at RT for 16 h. (TLC system: 70% Ethylacetate in Hexane, Rfvalue: 0.3). To the reaction mixture was added ice cold water (100 mL)and stirred for 15 min. The resultant solid was filtered, dried andpurified by combiflash chromatography (using silicagel column, elutedwith 3% Methanol in DCM) to afford(9S)-N2-cyclopropyl-N10-(pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(4.05 g, 10.70 mmol, 60.5% yield) as an off-white solid. LCMS (m/z):379.16 [M+H]⁺, Rt=1.97 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.70-14.14 (m, 1H), 8.29-8.42 (m, 1H),8.16 (dt, J=8.33, 0.88 Hz, 2H), 7.97 (d, J=7.89 Hz, 1H), 7.72 (td,J=7.84, 1.64 Hz, 1H), 7.58 (d, J=7.89 Hz, 1H), 7.04 (ddd, J=7.29, 4.99,1.10 Hz, 1H), 4.97 (t, J=2.19 Hz, 1H), 3.29-3.42 (m, 3H), 3.10 (ddt,J=10.93, 7.32, 3.92, 3.92 Hz, 1H), 2.96 (br d, J=13.59 Hz, 1H),2.16-2.29 (m, 1H), 1.86-1.99 (m, 1H), 1.59 (s, 1H), 1.32-1.45 (m, 2H),0.81-1.02 (m, 4H).

Example 37 Synthesis of(9S)-N10-(pyridin-2-yl)-N2-(2,2,2-trifluoroethyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid(7 g, 20.63 mmol) in DMF (50 mL), under nitrogen at 0° C. was addedDIPEA (10.81 mL, 61.9 mmol), HATU (15.69 g, 41.3 mmol) and2,2,2-trifluoroethanamine (2.248 g, 22.69 mmol) then the reaction wasstirred at RT for 16 h. (TLC system: 70% Ethylacetate in Hexane, Rfvalue: 0.3). To the reaction mixture was added ice cold water (200 mL)and stirred for 30 min. The resultant solid was filtered, dried andpurified by combiflash chromatography (using silicagel column, elutedwith 3% Methanol in DCM) to afford the product as an off-white solid.The solid product was dissolved in Ethanol (500 mL) at 80° C. and addedpalladium scavenger (Silicycle Brand, 8 g) and continued heating for 5h. The reaction mixture was filtered through celite, filtrate wasconcentrated to afford(9S)-N10-(pyridin-2-yl)-N2-(2,2,2-trifluoroethyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(8.1 g, 19.17 mmol, 93% yield) as an off-white solid. LCMS (m/z): 421.1[M+H]⁺, Rt=2.23 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 14.13-14.37 (m, 1H), 8.71 (br t, J=5.92Hz, 1H), 8.25 (dd, J=4.93, 0.99 Hz, 1H), 8.11 (d, J=8.33 Hz, 1H), 7.96(d, J=7.89 Hz, 1H), 7.70-7.77 (m, 1H), 7.61 (d, J=7.89 Hz, 1H),7.00-7.05 (m, 1H), 4.97 (t, J=2.30 Hz, 1H), 4.13-4.37 (m, 2H), 3.27-3.45(m, 3H), 2.97 (br d, J=13.59 Hz, 1H), 2.18-2.27 (m, 1H), 1.87-2.00 (m,1H), 1.34-1.46 (m, 2H).

Example 38 Synthesis of(9S)-3-chloro-N10-(pyridin-2-yl)-N2-(2,2,2-trifluoroethyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a suspension of(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (900 mg, 2.408 mmol) in Acetonitrile (15 mL), under nitrogen at RTwas added DIPEA (1.262 mL, 7.22 mmol), 2-chloro-1-methylpyridin-1-iumiodide (615 mg, 2.408 mmol) and 2,2,2-trifluoroethanamine (238 mg, 2.408mmol), then the resulting reaction mixture was stirred for 16 h. (TLCsystem: 75% EtOAc in Hexane, R_(f) value: 0.30). Reaction mass wasdiluted with 100 mL of water and extracted with EtOAc (2×150 mL). Thecombined organic layer was washed with brine (80 mL), dried over Na₂SO₄,filtered and concentrated to get crude. The crude material was purifiedby combiflash (using silica gel column, 70% EtOAc in Hexane). Fractionscontaining compound were combined and concentrated to afford the desiredcompound(9S)-3-chloro-N10-(pyridin-2-yl)-N2-(2,2,2-trifluoroethyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(52 mg, 0.114 mmol, 4.75% yield) as an off white solid. LCMS (m/z):455.05 [M+H]⁺, Rt=2.17 min. 111 NMR (400 MHz, CDCl₃): δ ppm 13.43 (s,1H), 8.29-8.49 (m, 1H), 8.25 (d, J=4.17 Hz, 1H), 8.11 (d, J=8.33 Hz,1H), 7.67-7.84 (m, 1H), 7.59 (s, 1H), 6.95-7.14 (m, 1H), 4.97 (br s,1H), 4.12-4.37 (m, 2H), 3.28-3.44 (m, 3H), 2.95 (br d, J=13.81 Hz, 1H),2.10-2.33 (m, 1H), 1.86-2.07 (m, 1H), 1.25-1.50 (m, 2H).

Example 39 Synthesis of(9S)-3-chloro-N10-(pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-3-chloro-10-(pyridin-2-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (500 mg, 1.338 mmol) in DMF (6 mL), under nitrogen at RT was addedDIPEA (0.701 mL, 4.01 mmol), HATU (1017 mg, 2.68 mmol) and(R)-1,1,1-trifluoropropan-2-amine (182 mg, 1.605 mmol), then theresulting reaction mixture was stirred at RT for 16 h. (TLC system: 5%MeOH in DCM, R_(f) value: 0.30). Reaction mass was diluted with 50 mL ofice cold water, extracted with EtOAc (2×100 mL). The combined organiclayer was washed with brine (80 mL), dried over Na₂SO₄, filtered andconcentrated to get crude compound. The crude material was purified bycombiflash (using silica gel column, 80% EtOAc in Hexane). Fractionscontaining compound were combined and concentrated to give the desiredcompound with 60% purity by LCMS. This was further purified by prep HPLC(Column: Kromsil phenyl (150×25) mm 10μ; MP-A: 10 mM AmmoniumBicarbonate (aq), MP-B: Acetonitrile; Method: 50:50; Flow: 20 ml/minSolubility: ACN+THF) to afford the desired product(9S)-3-chloro-N10-(pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(50 mg, 0.106 mmol, 7.93% yield) as an off white solid. LCMS (m/z):469.16 [M+H]⁺, Rt=2.22 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 12.89 (s, 1H), 8.27 (d, J=4.17 Hz, 1H),8.15 (d, J=8.33 Hz, 1H), 7.64-7.80 (m, 2H), 7.59 (s, 1H), 6.90-7.15 (m,1H), 5.09 (dq, J=16.83, 7.55 Hz, 1H), 4.89-5.01 (m, 1H), 3.26-3.44 (m,3H), 2.94 (br d, J=13.81 Hz, 1H), 2.14-2.35 (m, 1H), 1.85-2.09 (m, 1H),1.39-1.63 (m, 3H), 1.40-1.32 (s, 2H).

Example 40 Synthesis of(9S)-N10-(isoxazol-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-(isoxazol-3-ylcarbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (400 mg, 1.215 mmol) and (R)-1,1,1-trifluoropropan-2-amine (206 mg,1.822 mmol) in DMF (15 mL) under nitrogen at RT was added HATU (554 mg,1.458 mmol), DIPEA (0.424 mL, 2.429 mmol) and stirred for 16 h. (TLCsystem: Ethylacetae. Rf value: 0.6). The reaction mixture was quenchedwith ice cold water (30 mL) and extracted with ethyl acetate (3×50 mL).The combined organic layer was washed with brine solution (30 mL), driedover anhydrous sodium sulphate and concentrated under reduced pressureto obtain crude compound. The crude product was purified by flash columnchromatography (Silica gel, uding 60% Ethylacetate in petether) toafford the desired product(9S)-N10-(isoxazol-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(160 mg, 0.373 mmol, 30.7% yield) as an off white solid. LCMS (m/z):425.09 [M+H]⁺, Rt=2.14 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.55 (s, 1H), 8.30 (d, J=1.32 Hz, 1H),7.96 (d, J=7.89 Hz, 1H), 7.62 (d, J=7.89 Hz, 1H), 7.23 (br s, 1H), 6.99(d, J=1.75 Hz, 1H), 4.89-5.05 (m, 2H), 3.32-3.47 (m, 3H), 2.96 (br d,J=13.81 Hz, 1H), 2.22 (dt, J=14.69, 2.96 Hz, 1H), 1.90-2.02 (m, 1H),1.64 (d, J=7.02 Hz, 3H), 1.33-1.45 (m, 2H).

Example 41 Synthesis of(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-(2,2,2-trifluoroethyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (200 mg, 0.508 mmol) in N,N-Dimethylformamide (5 mL), HATU (193 mg,0.508 mmol) and DIPEA (0.089 mL, 0.508 mmol) were added under nitrogenatmosphere at 28° C. and stirred for 30 min. at room temperature andfollowed by 2,2,2-trifluoroethanamine hydrochloride (103 mg, 0.763 mmol)was added and the reaction mixture was stirred at 28° C. for 16 h. (TLC:10% MeOH/CH₂Cl₂, R_(f) value: 0.4). The reaction mixture was dilutedwith water (40 mL) and extracted with EtOAc (2×40 mL). The combinedorganic layer was dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to get crude compound. The crude product was purified byflash column chromatography (Silicagel: 100-200 Mesh, Eluent: 5%Methanol/DCM) to afford the desired product(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-(2,2,2-trifluoroethyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(100 mg, 0.211 mmol, 41.5% yield) as an off white solid. LCMS (m/z):475.10 [M+H]⁺, R_(t)=1.96 min.

¹H NMR (400 MHz, CDCl₃-d): δ ppm 13.85 (s, 1H), 10.39 (s, 1H), 8.75 (d,J=8.33 Hz, 1H), 8.04-7.90 (m, 2H), 7.63 (d, J=7.89 Hz, 1H), 7.03 (d,J=8.55 Hz, 1H), 5.00 (d, J=2.19 Hz, 1H), 4.37-4.12 (m, 2H), 3.48-3.30(m, 3H), 3.00 (d, J=13.59 Hz, 1H), 2.69 (s, 3H), 2.37-2.25 (m, 1H),2.07-1.88 (m, 1H), 1.48-1.39 (m, 2H).

Example 42 Synthesis of(9S)-N2-cyclopropyl-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (300 mg, 0.763 mmol), HATU (435 mg, 1.144 mmol) and DIPEA (0.400mL, 2.288 mmol) in DMF (15 mL) was added cyclopropanamine (65.3 mg,1.144 mmol) at room temperature and stirred the reaction mixture at RTfor 16 h. (TLC: neat ethyl acetate, R_(f) value: 0.3, UV active). Thereaction mixture was diluted with water (100 mL) and extracted withEtOAc (2×100 mL). The combined organic layer was washed with saturatedbrine solution (50 mL) and dried over anhydrous Na₂SO₄, filtered andfiltrate was evaporated to obtain crude compound. The crude product waspurified by flash column chromatography (Silica gel: 100-200 Mesh,Eluent: 80% ethyl acetate in n-hexane) to afford the desired product(9S)-N2-cyclopropyl-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(90 mg, 0.202 mmol, 26.5% yield) as an off white solid. LCMS (m/z):433.16 [M+H]⁺, R_(t)=1.76 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.67-13.63 (m, 1H), 13.65 (s, 1H), 11.16(s, 1H), 8.71 (d, J=8.33 Hz, 1H), 7.93 (d, J=7.89 Hz, 1H), 7.60 (d,J=8.11 Hz, 1H), 7.52 (d, J=2.85 Hz, 1H), 7.03 (d, J=8.55 Hz, 1H), 5.01(s, 1H), 3.42-3.31 (m, 3H), 3.12-3.05 (m, 1H), 3.00 (d, J=13.81 Hz, 1H),2.73 (s, 3H), 2.27 (d, J=14.47 Hz, 1H), 2.01-1.88 (m, 1H), 1.45-1.36 (m,2H), 0.91-0.85 (m, 3H).

Example 43 Synthesis of(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (200 mg, 0.508 mmol), HOBT (97 mg, 0.635 mmol), EDC (122 mg, 0.635mmol) and DIPEA (0.266 mL, 1.525 mmol) in DMF (15 mL) was added(R)-1,1,1-trifluoropropan-2-amine (69.0 mg, 0.610 mmol) at roomtemperature and stirred the reaction mixture at RT for 16 h. (TLC: neatethyl acetate, R_(f) value: 0.3, UV active). The reaction mixture wasdiluted with water (100 mL) and extracted with EtOAc (2×100 mL). Thecombined organic layer was washed with brine solution (50 mL) and driedover anhydrous Na₂SO₄, filtered and filtrate was evaporated to obtaincrude compound. The crude product was purified by flash columnchromatography (using 100-200 silica gel, compound eluted at 80% ethylacetate in n-hexane) to afford the desired product(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(119 mg, 0.237 mmol, 46.6% yield) as an off white solid. LCMS: (m/z):489.2 [M+H]⁺, Rt=2.06 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.63 (s, 1H), 10.14 (s, 1H), 8.69 (d,J=8.33 Hz, 1H), 7.96 (d, J=7.89 Hz, 1H), 7.69 (d, J=9.21 Hz, 1H), 7.62(d, J=7.89 Hz, 1H), 7.03 (d, J=8.55 Hz, 1H), 5.12-4.95 (m, 2H),3.51-3.18 (m, 3H), 2.93-3.05 (m, 1H), 2.68 (s, 3H), 2.29 (d, J=14.47 Hz,1H), 2.04-1.87 (m, 1H), 1.57 (s, 3H), 1.49-1.25 (m, 2H).

Example 44 Synthesis of(9S)-N10-(4-(2-methylthiazol-5-yl)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-N10-(4-bromopyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(350 mg, 0.682 mmol) and2-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)thiazole (230mg, 1.023 mmol) in 1,4-Dioxane (20 mL) and water (1.0 mL) under nitrogenat RT was added K₃PO₄ (434 mg, 2.046 mmol) and degassed by purging agonfor 20 min. Then added PdCl₂(dppf)-CH₂Cl₂ adduct (55.7 mg, 0.068 mmol)and the mixture was at 110° C. for 3.5 h. (TLC system: 100% Ethylacetae. Rf value: 0.3). The reaction mixture was cooled to RT anddiluted with water (30 mL), extracted with ethyl acetate (2×50 mL). Thecombined organic layer was washed with brine solution (50 mL) and driedover anhydrous sodium sulphate and concentrated under reduced pressureto obtain crude compound. The crude product was purified by flash columnchromatography (100-200 silicagel eluted with 50% of EtOAc in Pet ether)to afford the desired product(9S)-N10-(4-(2-methylthiazol-5-yl)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(320 mg, 0.594 mmol, 87% yield) as an off white solid. LCMS (m/z):532.20 [M+H]⁺, Rt=2.60 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.88 (s, 1H), 8.39 (s, 1H), 8.26 (d,J=5.26 Hz, 1H), 8.13-7.89 (m, 3H), 7.62 (d, J=8.11 Hz, 1H), 7.15 (dd,J=5.26, 1.53 Hz, 1H), 5.20-5.06 (m, 1H), 5.00 (br s, 1H), 3.44-3.21 (m,3H), 3.03-2.90 (m, 1H), 2.76 (s, 3H), 2.25 (br d, J=14.47 Hz, 1H),2.05-1.89 (m, 1H), 1.62 (d, J=7.02 Hz, 3H), 1.49-1.36 (m, 2H).

Example 45 Synthesis of(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-(1,1,1-trifluoro-3-hydroxypropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred suspension of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (4.5 g, 11.44 mmol) in pyridine (40 mL) under nitrogen at 0° C. wasadded EDC (4.39 g, 22.88 mmol) followed by2-amino-3,3,3-trifluoropropan-1-ol (2.215 g, 17.16 mmol) and thereaction mixture was stirred at RT for 16 h. (TLC system 5% Methanol inDCM. Rf value 0.3). The reaction mixture was concentrated and theresidue was dissolved in EtOAc (200 mL) and washed with water (2×100mL). Combined organic layer was dried over anhydrous Na₂SO₄, filteredand concentrated to get crude as brown solid. The solid was trituratedwith diethylether (50 mL), filtered and dried to get the desiredcompound as diastereomeric mixture. The diastereomers were separated bypreparative chiral SFC (Column/dimensions: Chiralpak AD-H (250×30) mm,5μ; % CO₂: 50.0; % Co-solvent: 50.0% (MeOH); Total Flow: 100.0 g/min,Back Pressure: 100.0 bar; UV: 213 nm, Stack time: 6.7 min, Load/inj:95.0 mg, Solubility: MeOH, Total No of injections: 60, Instrumentdetails: Make/Model: Thar SFC-200 NEW-1).

Peak-1 Collected fraction from SFC was concentrated and triturated withdiethylether (20 mL), dried and grinded in motor to afford(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-(1,1,1-trifluoro-3-hydroxypropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(920 mg, 1.821 mmol, 15.92% yield) as an off-white solid. LCMS (m/z):505.23 [M+H]⁺. Rt=1.85 min.

Example 46 Synthesis of(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-(1,1,1-trifluoro-3-hydroxypropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred suspension of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (4.5 g, 11.44 mmol) in pyridine (40 mL) under nitrogen at 0° C. wasadded EDC (4.39 g, 22.88 mmol) followed by2-amino-3,3,3-trifluoropropan-1-ol (2.215 g, 17.16 mmol) and thereaction mixture was stirred at RT for 16 h. (TLC system 5% Methanol inDCM. Rf value 0.3). The reaction mixture was concentrated and theresidue was dissolved in EtOAc (200 mL) and washed with water (2×100mL). Combined organic layer was dried over anhydrous Na₂SO₄, filteredand concentrated to get crude as brown solid. The solid was trituratedwith diethylether (50 mL), filtered and dried to get the desiredcompound as diastereomeric mixture. The diastereomers were separated bypreparative chiral SFC (Column/dimensions: Chiralpak AD-H (250×30) mm,5μ; % CO₂: 50.0; % Co-solvent: 50.0% (MeOH); Total Flow: 100.0 g/min,Back Pressure: 100.0 bar; UV: 213 nm, Stack time: 6.7 min, Load/inj:95.0 mg, Solubility: MeOH, Total No of injections: 60, Instrumentdetails: Make/Model: Thar SFC-200 NEW-1).

Peak-2 Collected fraction from SFC was concentrated and washed withdiethylether (20 mL), dried and grinded in motor to afford(9S)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-(1,1,1-trifluoro-3-hydroxypropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(1 g, 1.966 mmol, 17.19% yield) as an off-white solid. LCMS (m/z):505.2323 [M+H]⁺. Rt=1.85 min

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.15 (s, 1H), 12.92-13.08 (m, 1H),8.56 (d, J=8.99 Hz, 1H), 8.39 (d, J=8.33 Hz, 1H), 7.60-7.85 (m, 2H),7.03 (d, J=8.55 Hz, 1H), 5.24 (t, J=6.36 Hz, 1H), 4.74-4.91 (m, 2H),3.85 (t, J=6.25 Hz, 2H), 3.43 (dd, J=13.48, 1.64 Hz, 3H), 2.81-3.04 (m,1H), 2.57 (s, 3H), 2.05 (br d, J=18.42 Hz, 2H), 1.33 (br d, J=8.33 Hz,2H).

Example 47 Synthesis of(9S)-4-methyl-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-4-methyl-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxamide(300 mg, 0.914 mmol) in THF (30 mL) at RT was added TEA (0.764 mL, 5.48mmol), triphosgene (271 mg, 0.914 mmol) and stirred for 30 min, thenadded 6-methyl-1H-pyrazolo[3,4-b]pyridin-3-amine (271 mg, 1.827 mmol)and the reaction mixture was heated at 65° C. for 16 h. (TLC elutingsystem: 100% EtOAc: R_(f)-0.4; UV active). The reaction mixture wascooled to RT, quenched with water (20 mL) and extracted into EtOAc (2×35mL). Organic layer was separated, dried over anhydrous sodium sulphate,filtered and filtrate was evaporated to get the crude product. The crudewas purified by chromatography (GRACE using C-18 reserval column, Mobilephase A: 0.1% Formic Acid in water; B: ACN, eluent 44% B in A) andcombined fractions were concentrated and basified with saturated NaHCO₃.The aqueous layer was extracted with DCM, combined DCM layer was driedover anhydrous Na₂SO₄, filtered and filtrate was evaporated to afford(9S)-4-methyl-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(105 mg, 0.204 mmol, 22.34% yield) as yellow solid. LCMS (m/z): 503.25[M+H]⁺, R_(t)=2.38 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.87 (s, 1H), 9.81 (br s, 1H), 8.70 (d,J=8.1 Hz, 1H), 7.84 (s, 1H), 7.68 (br d, J=8.8 Hz, 1H), 7.02 (d, J=8.3Hz, 1H), 5.11-4.90 (m, 2H), 3.39 (br d, J=13.4 Hz, 1H), 3.21 (br d,J=6.8 Hz, 2H), 2.97 (br d, J=14.0 Hz, 1H), 2.66 (s, 3H), 2.43 (s, 3H),2.38-2.13 (m, 1H), 2.13-1.86 (m, 1H), 1.63-1.49 (m, 3H), 1.42 (br d,J=6.1 Hz, 2H).

Example 48 Synthesis of(9S)-N2-(2,2-difluorocyclopropyl)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (500 mg, 2.54 mmol) in DMF (10 mL) stirred under nitrogen at roomtemp were added HATU (2.9 g, 7.63 mmol) and DMAP (0.621 g, 5.08 mmol).To this 2,2-difluorocyclopropanamine, Hydrochloride (0.823 g, 6.35 mmol)was added and the reaction mixture was stirred at 50° C. for 16 h (TLCeluent: 10% MeOH in DCM: R_(f)-0.4; UV active). Reaction mixture wasallowed to cool to room temperature and diluted with ice water,extracted with ethylacetate (2×50 mL). The combined organic layer waswashed with brine solution and dried over anhydrous sodium sulphate andconcentrated under reduced pressure to afford crude compound. The crudeproduct was purified by column chromatography (silicagel: 100-200 Mesh,Eluent: 2% Methanol in DCM) to afford the diastereomeric mixture andsubmitted for SFC separation (Conditions: (Column/dimensions): ChiralpakIC (250×30) mm, 5 μ% CO₂: 50.0% Co solvent: 50.0% (100% M ETHANOL),Total Flow: 100.0 g/min, Back Pressure: 100.0 bar, UV: 217 nm, Stacktime: 16.5 min, Load/inj: 46.0 mg, Solubility: Methanol, Total No ofinjections: 7, Instrument details: Make/Model: Thar SFC-200 (OLD)) toafford two peaks as peak-I and peak-II.

Peak-II:(9S)-N2-(2,2-difluorocyclopropyl)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(73 mg, 0.155 mmol, 6.08% yield) as a pale brown solid. LCMS (m/z):469.2 [M+H]⁺, Rt=1.94 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.2 (s, 1H), 12.8 (s, 1H), 8.64 (br s,1H), 8.41 (d, J=8.55 Hz, 1H), 7.74-7.6 (m, 1H), 7.64-7.62 (m, 1H), 7.05(d, J=8.55 Hz, 1H), 4.87 (s, 1H), 3.56-3.39 (m, 2H), 3.35 (br s, 2H),2.87 (d, J=13.37 Hz, 1H), 2.55 (s, 3H), 2.07-1.9 (m, 4H), 1.35-1.22 (m,2H).

Example 49 Synthesis of(9S)-N10-(5-((R)-2,3-dihydroxypropoxy)pyrazin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N10-(5-((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(200 mg, 0.354 mmol) in Methanol (10 mL) under nitrogen atmosphere wasadded aq HCl (4 ml, 16.00 mmol) at 0° C. The resulted reaction mixturewas stirred at 28° C. for 1 h. (TLC System: 5% Methanol in DCM, R_(f):0.3, UV active). The solvent was evaporated under reduced pressure,basified with saturated NaHCO₃ solution (till pH: 8-9) and theprecipitated solid was filtered to afford the desired product(9S)-N10-(5-((R)-2,3-dihydroxypropoxy)pyrazin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(110 mg, 0.207 mmol, 58.4% yield) as an off-white solid. LCMS (m/z):526.26 [M+H]⁺, R_(t)=1.87 min.

¹H NMR (400 MHz, CDCl₃): δ 13.64 (s, 1H), 8.99 (s, 1H), 8.96-7.96 (m,2H), 7.71 (d, J=9.43 Hz, 1H), 7.62 (d, J=7.89 Hz, 1H), 5.15-4.93 (m,2H), 4.54-4.39 (m, 2H), 4.16-4.03 (m, 1H), 3.85-3.66 (m, 2H), 3.46-3.27(m, 3H), 3.08 (d, J=3.95 Hz, 1H), 2.96 (d, J=14.03 Hz, 1H), 2.33-2.18(m, 2H), 2.02-1.87 (m, 1H), 1.57 (d, J=7.02 Hz, 3H), 1.42 (s, 2H).

Example 50 Synthesis of(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide (270 mg, 0.496 mmol) in Methanol (5 mL) at0° C. was added hydrochloric acid (0.030 mL, 0.992 mmol) drop wise overa period of 5 min. Then the reaction mixture was stirred at RT for 2 h.(TLC eluent: 5% MeOH in DCM: R_(f)-0.1; UV active). The reaction mixturewas neutralization with sodium bicarbonate solution and filtered theobtain solid, washed with diethyl ether (2×30 ml), n-pentane (2×30 ml)to afford the desired product(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(150 mg, 0.297 mmol, 59.8% yield) as an off white solid. LCMS (m/z):505.23 [M+H]⁺, Rt=1.50 min.

¹H NMR (400 MHz, TFA-d): δ ppm 8.58-8.49 (m, 1H), 8.28 (d, J=7.7 Hz,1H), 8.01-7.98 (m, 1H), 7.39-7.20 (m, 2H), 5.40-5.36 (m, 1H), 4.68-4.39(m, 3H), 4.45-3.80 (m, 6H), 3.62-3.49 (m, 1H), 2.39-2.29 (m, 1H),2.30-1.80 (m, 5H).

Example 51 Synthesis of(9S)-3-chloro-N10-(4-(2-methyloxazol-5-yl)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a solution of(9S)-3-chloro-N-((R)-1,1,1-trifluoropropan-2-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxamide(700 mg, 2.007 mmol) in THF (20 mL) at RT, was added TEA (1.399 mL,10.04 mmol) followed by triphosgene (357 mg, 1.204 mmol) and stirred for30 min. then added 4-(2-methyloxazol-5-yl)pyridin-2-amine (527 mg, 3.01mmol) and the reaction mixture was heated to 80° C. for 16 h. (TLCsystem: 100% Ethylacetate, Rf value: 0.4). The reaction mixture dilutedwith cold water (50 mL) and extracted with EtOAc (2×50 mL). The combinedorganic layer was washed with brine solution (30 mL), dried overanhydrous sodiumsulphate and concentrated under reduced pressure toobtain crude compound. The crude product was purified by prep HPLC(Column: Kromosil Phenyl C18 (150*25) mm 10 u, Mobile Phase-A: 10 mMAmmonium Bicarbonate (Aq), Mobile Phase-B: Acetonitrile, Gradient: TimeT/% B=0/45, 12/45, 12.5/100, 15/100, 15.5/45, Column Temp: Ambient, FlowRate: 30 ml/min, Diluent: Acetonitrile+THF+Water) to afford(9S)-3-chloro-N10-(4-(2-methyloxazol-5-yl)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido-[2,3-b][1,4]-diazocine-2,10(7H)-dicarboxamide(95 mg, 0.171 mmol, 8.53% yield) as an off white solid. LCMS (m/z):550.20 [M+H]⁺, Rt=2.36 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.04 (s, 1H), 9.22 (d, J=8.77 Hz, 1H),8.36 (s, 1H), 8.30 (d, J=5.26 Hz, 1H), 7.81 (d, J=12.50 Hz, 2H), 7.39(dd, J=5.26, 1.53 Hz, 1H), 4.93-4.71 (m, 2H), 3.48-3.39 (m, 1H),3.32-3.30 (m, 2H), 2.88 (br d, J=13.81 Hz, 1H), 2.53 (s, 3H), 2.02-1.88(m, 2H), 1.46 (d, J=7.02 Hz, 3H), 1.38-1.22 (m, 2H).

Example 52 Synthesis of(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(230 mg, 0.422 mmol) in Methanol (5 mL) at 0° C. was added hydrochloricacid (0.026 mL, 0.845 mmol), drop wise over a period of 5 min. Then thereaction mixture was stirred at RT for 2 h. (TLC eluent: 5% MeOH in DCM:0.1; UV active). Then reaction mixture was neutralized with sodiumbicarbonate solution and obtained solid was filtered and washed withether (2×30 ml), pentane (2×30 ml) to afford the desired product(9S)-N2-(2,2-difluorocyclopropyl)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(175 mg, 0.346 mmol, 82% yield) as an off white solid. LCMS (m/z):505.23 [M+H]⁺, Rt=1.50 min.

¹H NMR (400 MHz, TFA-d): 6 ppm 8.56-8.50 (m, 1H), 8.39 (d, J=7.7 Hz,1H), 8.02 (m, 1H), 7.24-7.10 (m, 2H), 5.41-5.39 (m, 1H), 4.64-4.41 (m,3H), 4.39-3.89 (m, 6H), 3.68-3.51-3.39 (m, 1H), 2.41-2.29 (m, 1H),2.22-1.76 (m, 5H).

Example 53 Synthesis of(9S)-N2-(2,2-difluorocyclopropyl)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (500 mg, 2.54 mmol) in DMF (10 mL) stirred under nitrogen at roomtemp were added HATU (2.9 g, 7.63 mmol) and DMAP (0.621 g, 5.08 mmol).To this 2,2-difluorocyclopropanamine, Hydrochloride (0.823 g, 6.35 mmol)was added and the reaction mixture was stirred at 50° C. for 16 h (TLCeluent: 10% MeOH in DCM: R_(f)-0.4; UV active). Reaction mixture wasallowed to cool to room temperature and diluted with ice water,extracted with ethylacetate (2×50 mL). The combined organic layer waswashed with brine solution and dried over anhydrous sodium sulphate andconcentrated under reduced pressure to afford crude compound. The crudeproduct was purified by column chromatography (silicagel: 100-200 Mesh,Eluent: 2% Methanol in DCM) to afford the diastereomeric mixture andsubmitted for SFC separation (Conditions: (Column/dimensions): ChiralpakIC (250×30) mm, 5 μ% CO₂: 50.0% Co solvent: 50.0% (100% M ETHANOL),Total Flow: 100.0 g/min, Back Pressure: 100.0 bar, UV: 217 nm, Stacktime: 16.5 min, Load/inj: 46.0 mg, Solubility: Methanol, Total No ofinjections: 7, Instrument details: Make/Model: Thar SFC-200 (OLD)) toafford two peaks as peak-I and peak-II.

Peak-I:(9S)-N2-(2,2-difluorocyclopropyl)-N10-(6-methyl-1H-pyrazolo[3,4-b]pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(70 mg, 0.147 mmol, 5.80% yield) as an off white solid. LCMS (m/z):469.2 [M+H]⁺, Rt=1.92 min.

NMR (400 MHz, DMSO-d₆): δ ppm 13.26 (s, 1H), 12.99 (s, 1H), 8.64 (br s,1H), 8.41 (d, J=8.55 Hz, 1H), 7.74-7.69 (m, 1H), 7.66-7.62 (m, 1H), 7.03(d, J=8.55 Hz, 1H), 4.87 (br s, 1H), 3.54-3.39 (m, 2H), 3.32 (br s, 2H),2.88 (d, J=13.37 Hz, 1H), 2.57 (s, 3H), 2.07-1.88 (m, 4H), 1.35-1.24 (m,2H).

Example 54 Synthesis of(9S)-N10-(5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-10-((5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)carbamoyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2-carboxylicacid (300 mg, 0.699 mmol) and HATU (398 mg, 1.048 mmol) inN,N-Dimethylformamide (10 mL) was added DIPEA (0.488 mL, 2.79 mmol) andfollowed by (R)-1,1,1-trifluoropropan-2-amine Hydrochloride (104 mg,0.699 mmol) at room temperature to the reaction mixture. The resultedreaction mixture was stirred for 16 h. at 28° C. (TLC eluent: 10% MeOHin DCM: R_(f)-0.3; UV active). The reaction mixture was poured in towater (30 mL) and extracted with EtOAc (2×20 mL). The combined organiclayer was washed with brine solution dried over Na₂SO₄, filtered andevaporated to get crude compound. The crude compound was purified byGRACE (C-18 reserval column, Eluent: 60% of MeOH and 0.1% of Formic Acidin water) to afford the desired product(9S)-N10-(5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(169 mg, 0.310 mmol, 44.4% yield) as a pale yellow solid. LCMS (m/z):525.25 [M+H]⁺, R_(t)=1.86 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.62 (s, 1H), 8.14 (d, J=8.99 Hz, 1H),8.06-7.90 (m, 3H), 7.71-7.51 (m, 1H), 7.41-7.24 (m, 1H), 5.02-5.21 (m,1H), 4.98 (br s, 1H), 4.24-4.03 (m, 3H), 3.95-3.83 (m, 1H), 3.76-3.82(m, 1H), 3.41-3.27 (m, 3H), 2.95 (d, J=14.03 Hz, 1H), 2.55 (br s, 1H),2.23 (d, J=15.13 Hz, 1H), 1.87-2.01 (m, 2H), 1.61 (d, J=7.02 Hz, 3H),1.48-1.34 (m, 2H).

Example 55 Synthesis of(9S)-N10-(6-((R)-2,3-dihydroxypropoxy)pyridazin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N10-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridazin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(220 mg, 0.389 mmol) in Methanol (10 mL) under nitrogen was added Aq HCl(0.8 ml, 3.20 mmol) at 0° C. The reaction mixture was stirred at 28° C.for 1 h. (TLC System: 5% Methanol in DCM, Rf: 0.3, UV active). Thesolvent was evaporated under reduced pressure, basified with saturatedNaHCO₃ solution (till pH: 8-9) and extracted with DCM (3×30 mL). Thecombined organic layer was washed with water, brine solution, filteredand evaporated to get crude compound. The crude compound was purified byPrep HPLC (conditions: MP-A: 10 Mm Ammonium Acetate (Aq) MP-B:Acetonitrile Column: Kromasil C18(250*21.2) mm, 10μ Method (T/% B):65:35 Flow: 20 ml/min Solubility: Acetonitrile+MeOH+THF) to afford thedesired product(9S)-N10-(6-((R)-2,3-dihydroxypropoxy)pyridazin-3-yl)-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide (130 mg, 0.234 mmol, 60.1% yield) as anoff-white solid. LCMS (m/z): 524.2 [M+H]⁺, R_(t)=5.98 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 14.34 (s, 1H), 8.42 (d, J=9.43 Hz, 1H),8.07 (d, J=9.21 Hz, 1H), 8.01 (d, J=7.89 Hz, 1H), 7.63 (d, J=7.89 Hz,1H), 7.10 (d, J=9.43 Hz, 1H), 5.12-5.00 (m, 1H), 4.96 (br s, 1H),4.65-4.54 (m, 2H), 4.16-4.08 (m, 1H), 3.82-3.66 (m, 2H), 3.49 (br s,1H), 3.43-3.30 (m, 3H), 2.97 (d, J=13.81 Hz, 1H), 2.34 (br s, 1H),2.26-2.17 (m, 1H), 2.01-1.89 (m, 1H), 1.71 (d, J=7.02 Hz, 3H), 1.47-1.36(m, 2H).

Example 56 Synthesis of(9S)-N10-(5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N10-(5-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(300 mg, 0.519 mmol) in methanol (15 mL) at 0° C. was added aq.HCl (1.5ml, 49.4 mmol) and stirred under nitrogen at RT for 1 h. (TLC eluent: 5%MeOH in DCM R_(f)-0.2; UV active). The reaction mixture at 0° C. wasbasified with saturated sodiumbicarbonate solution (till pH-8-9) andsolvent was evaporated under reduced pressure. The residue was dilutedwith water and resultant solid was filtered through Buchner funnel,dried under reduced pressure to afford crude compound. The crude waspurified by chromatography (GRACE using C-18 reserval column, Mobilephase A: 0.1% Formic Acid in water; B: ACN, eluent 45-50% B in A) andcombined fractions were concentrated then basified with saturatedNaHCO₃. The precipitated solid was filtered and dried to afford(9S)-N10-(5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(214 mg, 0.395 mmol, 76% yield) as an off white solid. LCMS (m/z):539.29 [M+H]⁺, R_(t)=2.14 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.83 (s, 1H), 8.14 (d, J=8.99 Hz, 1H),7.93-8.01 (m, 2H), 7.87 (s, 1H), 7.32 (dd, J=9.10, 2.96 Hz, 1H),5.04-5.17 (m, 1H), 4.95 (br s, 1H), 4.04-4.19 (m, 3H), 3.72-3.92 (m,2H), 3.37 (dd, J=13.59, 1.75 Hz, 1H), 3.14-3.25 (m, 2H), 2.92 (br d,J=13.37 Hz, 1H), 2.58 (d, J=4.60 Hz, 1H), 2.41 (s, 3H), 2.26 (br d,J=14.47 Hz, 1H), 1.86-2.02 (m, 2H), 1.53-1.64 (m, 3H), 1.33-1.47 (m,2H).

Example 57 Synthesis of(9S)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide

To a stirred solution of(9S)-N10-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(300 mg, 0.519 mmol) in methanol (15 mL) at 0° C. was added aq.HCl (1.5mL, 18.00 mmol) and stirred under nitrogen at RT for 1 h. (TLC eluent:5% MeOH in DCM R_(f)-0.3; UV active). The reaction mixture at 0° C. wasbasified with saturated sodiumbicarbonate solution (till pH-8-9) and thesolvent was evaporated under reduced pressure. The residue was dilutedwith water and the precipitated solid was filtered, dried under reducedpressure to afford crude. The crude was purified by chromatography(GRACE using C-18 reserval column, Mobile phase A: 0.1% Formic Acid inwater; B: ACN, eluent 50-55% B in A) and combined fractions wereconcentrated then basified with saturated NaHCO₃ solution. Theprecipitated solid was filtered and dried to afford(9S)-N10-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-4-methyl-N2-((R)-1,1,1-trifluoropropan-2-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-2,10(7H)-dicarboxamide(230 mg, 0.424 mmol, 82% yield) as off white solid. LCMS (m/z): 539.32[M+H]⁺, R_(t)=1.78 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 14.06 (s, 1H), 8.15 (br d, J=9.21 Hz,1H), 8.08 (d, J=5.92 Hz, 1H), 7.89 (s, 1H), 7.81 (d, J=2.41 Hz, 1H),6.59 (dd, J=5.70, 2.41 Hz, 1H), 5.00-5.15 (m, 1H), 4.93 (br s, 1H),4.10-4.22 (m, 3H), 3.69-3.90 (m, 2H), 3.37 (dd, J=13.48, 1.64 Hz, 1H),3.13-3.23 (m, 2H), 2.93 (br d, J=13.37 Hz, 1H), 2.57 (d, J=3.73 Hz, 1H),2.41 (s, 3H), 2.26 (br d, J=14.25 Hz, 1H), 1.86-2.07 (m, 2H), 1.50-1.63(m, 3H), 1.32-1.45 (m, 2H).

Example 58 Synthesis of(9S)-N-(5-fluoropyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-II from the intermediate SFC Separation, 0.700 g, 2.145 mmol) intetrahydrofuran (THF) (40 mL) stirred under nitrogen at room temperaturewas added triethylamine (1.794 mL, 12.87 mmol) and triphosgene (0.636 g,2.145 mmol). And then the reaction mixture was stirred at roomtemperature for 30 minutes. 5-fluoropyridin-2-amine (0.721 g, 6.43 mmol)was added at room temperature and then the reaction mixture was stirredat 65° C. for 16 hr. The reaction mixture was cooled to 28° C. andconcentrated to dryness. The residue was partitioned between water (10mL) and Dichloromethane (50 mL). Organic layer was separated and wasdried over anhydrous Na₂SO₄, filtered and filtrate was evaporated togive crude as brown solid (TLC eluent: 100% EtOAc: R_(f)-0.3; UVactive). The crude was purified by column chromatography using neutralalumina and was eluted with 35-40% EtOAc in Hexane to afford pure(9S)-N-(5-fluoropyridin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide(0.220 g, 0.468 mmol, 21.82% yield) as a off-white solid, LCMS (m/z):4653 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 12.14 (s, 1H), 8.41 (dd, J=9.2, 4.2 Hz, 1H),8.15 (d, J=3.0 Hz, 1H), 8.02 (s, 1H), 7.42 (ddd, J=9.2, 7.7, 3.0 Hz,1H), 5.51 (d, J=4.6 Hz, 1H), 3.76 (s, 1H), 3.58-3.46 (m, 1H), 3.41-3.17(m, 4H), 3.03 (s, 1H), 2.88-2.72 (m, 3H), 2.14 (d, J=13.7 Hz, 1H), 2.08(s, 1H), 2.00-1.77 (m, 3H), 1.53-1.42 (m, 2H), 1.37-1.21 (m, 2H).

Example 59 Synthesis of(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide

To a suspension of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-I from the intermediate SFC Separation, 500 mg, 1.532 mmol) intetrahydrofuran (50 mL) stirred at room temp for 10 min, triethylamine(0.641 mL, 4.60 mmol) and triphosgene (227 mg, 0.766 mmol) added. Thereaction mixture was stirred for 30 min pyrazin-2-amine (437 mg, 4.60mmol) was added. The reaction mixture was stirred for 16 hr at 65° C.The reaction mass concentrated and the residue was diluted with water(25 mL) and extracted with EtOAc (2×75 mL). The combined organic layerwas washed with water and saturated brine solution and dried overanhydrous Na₂SO₄, filtered and concentrated to give the crude as whitesolid (TLC eluent: 100% Ethyl Acetate: R_(f)-0.4; UV active). The crudeproduct was purified by column chromatography (neutral alumina) productwas eluted with 60% ethyl acetate in hexane to afford(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide(150 mg, 0.319 mmol, 20.82% yield) as a off white solid, LCMS (m/z):448.35 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 12.31 (s, 1H), 9.70 (d, J=1.5 Hz, 1H), 8.30(dd, J=2.5, 0.4 Hz, 1H), 8.26 (dd, J=2.6, 1.5 Hz, 1H), 8.03 (d, J=0.7Hz, 1H), 5.55 (d, J=4.6 Hz, 1H), 3.76 (s, 1H), 3.56-3.39 (m, 1H),3.29-3.20 (m, 4H), 3.02 (t, J=11.7 Hz, 1H), 2.93-2.78 (m, 3H), 2.15 (d,J=13.1 Hz, 1H), 2.06-1.82 (m, 4H), 1.54-1.42 (m, 2H), 1.36 (s, 1H).

Example 60 Synthesis of(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-II from intermediate SFC separation, 0.550 g, 1.685 mmol) inTetrahydrofuran (THF) (30 mL) stirred under nitrogen at room temperaturewas added Triethylamine (1.409 mL, 10.11 mmol) and Triphosgene (0.500 g,1.685 mmol). Then reaction mixture was stirred at room temperature for30 minute and then pyridin-3-amine (0.476 g, 5.06 mmol) was added atroom temperature. Then the reaction mixture was stirred at 65° C. for 16hr before being cooled to 28° C. and concentrated under reducedpressure. The residue was partitioned between water (10 mL) andDichloromethane (50 mL). Organic layer was separated and was dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude asbrown solid (TLC eluent: 100% EtOAc: R_(f)-0.3; UV active). The crudewas purified by column chromatography using neutral alumina and waseluted with 50-60% EtOAc in Hexane to afford pure(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide(0.264 g, 0.587 mmol, 34.8% yield) as a off-white solid, LCMS (m/z):447.3 [M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 11.74 (s, 1H), 8.59 (d, J=2.6 Hz, 1H), 8.44(ddd, J=8.4, 2.7, 1.5 Hz, 1H), 8.32 (dd, J=4.7, 1.6 Hz, 1H), 8.02 (d,J=0.8 Hz, 1H), 7.30 (dd, J=8.4, 4.7 Hz, 1H), 5.53 (d, J=5.0 Hz, 1H),3.76 (s, 1H), 3.58-3.41 (m, 1H), 3.24 (ddt, J=10.1, 7.3, 2.2 Hz, 4H),3.12 (t, J=11.7 Hz, 1H), 2.90-2.70 (m, 2H), 2.55 (dtt, J=15.5, 7.5, 3.9Hz, 1H), 2.12 (d, J=13.0 Hz, 1H), 2.01-1.84 (m, 3H), 1.81-1.66 (m, 1H),1.53 (td, J=12.7, 4.3 Hz, 2H), 1.35 (s, 1H), 1.26 (s, 1H).

Example 61 Synthesis of(9S)-N-(pyridin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide

To a suspension of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-I of the intermediate SFC separation, 600 mg, 1.840 mmol) intetrahydrofuran (50 mL) stirred at room temp for 10 min, triethylamine(0.796 mL, 5.521 mmol) and triphosgene (279 mg, 0.9202 mmol) added. Thereaction mixture was stirred for 30 min pyridin-2-amine (519 mg, 5.521mmol) was added. The reaction mixture was stirred for 16 hr at 65° C.The reaction mass concentrated and the residue was diluted with water(25 mL) and extracted with EtOAc (2×75 mL). The combined organic layerwas washed with water and saturated brine solution and dried overanhydrous Na₂SO₄, filtered and concentrated to give the crude as whitesolid (TLC eluent: 100% Ethyl Acetate: R_(f)-0.4; UV active). The crudeproduct was purified by column chromatography (neutral alumina) productwas eluted with 60% ethyl acetate in hexane to afford(9S)-N-(pyradin-3-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide(354 mg, 0.791 mmol, 43% yield) as a off white solid, LCMS (m/z): 447.28[M+H]⁺.

¹H NMR (400 MHz, CDCl₃): δ 11.66 (s, 1H), 8.60 (s, 1H), 8.47-8.40 (m,1H), 8.33 (s, 1H), 8.01 (s, J=0.66 Hz, 1H), 7.34-7.28 (m, 1H), 5.53 (brd, J=4.17 Hz, 1H), 3.76 (s, 1H), 3.47 (dt, J=12.17, 1.70 Hz, 1H),3.30-3.19 (m, 4H), 3.09 (t, J=11.62 Hz, 1H), 2.95-2.76 (m, 2H),2.65-2.48 (m, 1H), 2.18-2.07 (m, 1H), 1.98-1.85 (m, 3H), 1.78-1.68 (m,2H), 1.62-1.44 (m, 2H), 1.39-1.30 (m, 1H).

Example 62 Synthesis of(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide

To a solution of(9S)-2-(3-(trifluoromethyl)piperidin-1-yl)-′7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(Peak-II from intermediate SFC separation, 0.700 g, 2.145 mmol) inTetrahydrofuran (THF) (30 mL) stirred under nitrogen at room temperaturewas added Triethylamine (1.794 mL, 12.87 mmol) and Triphosgene (0.636 g,2.145 mmol). Then the reaction mixture was stirred at room temperaturefor 30 minutes before pyrazin-2-amine (0.612 g, 6.43 mmol) was added.Then the reaction mixture was stirred at 65° C. for 16 hr. The reactionmixture was cooled to 28° C. and concentrated under reduced pressure.The resulting residue was partitioned between water (10 mL) andDichloromethane (50 mL). Organic layer was separated and was dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to give crude asbrown solid (TLC eluent: 100% EtOAc: R_(f)-0.3; UV active). The crudewas purified by column chromatography using neutral alumina and waseluted with 50-60% EtOAc in Hexane to afford pure(9S)-N-(pyrazin-2-yl)-2-(3-(trifluoromethyl)piperidin-1-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-3-carboxamide(105 mg, 0.227 mmol, 10.58% yield) as a pale yellow solid, LCMS (m/z):448.3 [M+H]⁺.

¹H-NMR (400 MHz, CDCl₃): δ ppm 12.36 (s, 1H), 9.69 (d, J=1.53 Hz, 1H),8.28 (d, J=2.41 Hz, 1H), 8.23-8.25 (m, 1H), 8.03 (s, 1H), 5.54 (br d,J=4.17 Hz, 1H), 3.75 (br s, 1H), 3.48 (br d, J=10.30 Hz, 1H), 3.18-3.27(m, 4H), 3.05 (t, J=11.84 Hz, 1H), 2.74-2.85 (m, 3H), 2.14 (br d,J=12.93 Hz, 1H), 2.02 (dt, J=13.21, 4.03 Hz, 1H), 1.81-1.92 (m, 3H),1.39-1.52 (m, 2H), 1.27-1.36 (m, 1H).

Example 63 Synthesis of(9S)-3-chloro-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-2-(3-(trifluoromethyl)phenyl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.413 mmol) in THF (10 mL) was added NaH (33.9 mg, 1.413 mmol)at 0° C. and stirred under nitrogen atmosphere for 30 min. Then3-(pyridin-2-yl)-2H-pyrido[1,2-a][1,3,5]triazine-2,4(3H)-dione (340 mg,1.413 mmol) was added to this reaction mixture and stirred at 80° C. for16 h. (TLC eluent: Neat ethyl acetate; R_(f): 0.25). Reaction mixturewas quenched with ice water and extracted with Ethyl acetate (2×15 mL).The combined organic layer was washed with brine solution (20 mL) anddried over anhydrous sodium sulphate and concentrated under reducedpressure to obtain crude compound. The crude product was purified byflash column chromatograthy (silica-gel: 100-200 mesh, eluent: 70% Ethylacetate in petether) to afford the desired product(9S)-3-chloro-N-(pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(260 mg, 0.548 mmol, 38.7% yield) as an off white solid. LCMS (m/z):474.05 [M+H]⁺, R_(t)=3.0 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.40 (s, 1H), 8.30-8.23 (m, 2H),8.20-8.26 (m, 1H), 8.20-8.13 (m, 1H), 7.78-7.69 (m, 1H), 7.59-7.57 (m,2H), 7.52-7.48 (m, 1H), 6.97-6.91 (m, 1H), 5.02-4.97 (m, 1H), 3.41-3.37(m, 3H), 2.96-2.95 (m, 1H), 2.92 (d, J=14.03 Hz, 1H), 2.41-2.06 (m, 1H),1.63-1.21 (m, 2H).

Example 64 Synthesis of(9S)-3-chloro-N-(6-(((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.414 mmol) in methanol (10 mL) was added aq.HCl (0.345 mL,4.14 mmol, 36%) 0° C. and stirred at RT for 4 h. After completion of thereaction by TLC, the volatiles were evaporated under reduced pressure toget the crude (TLC eluent system: 100% EtOAc, Rf-0.5, UV active). Thecrude was diluted with water (5 ml) and basified with the 10% sodiumbicarbonate solution (up to pH 8), the precipitated solid was filtered,washed with the water and dried under vacuum to afford(9S)-3-chloro-N-(6-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(162 mg, 0.285 mmol, 68.8% yield) as an off white solid. LCMS (m/z):564.13 [M+H]⁺, R_(t)=2.50 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.10 (br s, 1H), 7.89-8.16 (m, 2H),7.72-7.80 (m, 1H), 7.63-7.70 (m, 1H), 7.47-7.61 (m, 2H), 6.42 (d, J=7.89Hz, 1H), 5.00 (br s, 1H), 3.92 (qd, J=11.91, 4.60 Hz, 2H), 3.67-3.81 (m,1H), 3.48 (br s, 2H), 3.27-3.42 (m, 2H), 2.98 (br d, J=12.93 Hz, 2H),2.80 (br d, J=5.48 Hz, 1H), 2.25 (br d, J=15.13 Hz, 2H), 1.94 (br d,J=12.50 Hz, 2H), 1.22-1.49 (m, 2H).

Example 65 Synthesis of(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(270 mg, 0.447 mmol), in methanol (10 mL) was added aq.HCl (1 mL, 32.9mmol, 36%) 0° C. and stirred at RT for 4 h. After completion of thereaction, the volatiles were evaporated under reduced pressure to getthe crude (TLC eluent system: 100% EtOAc, Rf-0.5, UV active). The crudewas diluted with the water (5 ml) and basified with the 10% sodiumbicaronate solution, the precipitated solid was filtered and was washedwith the water and dried under vacuum to afford(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(142 mg, 0.248 mmol, 55.5% yield) as an off white solid. LCMS (m/z):564.13 [M+H]⁺, R_(t)=2.51 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.11 (s, 1H), 7.89-8.14 (m, 2H),7.61-7.82 (m, 2H), 7.44-7.61 (m, 3H), 6.42 (d, J=7.89 Hz, 1H), 5.00 (brs, 1H), 3.83-4.04 (m, 2H), 3.63-3.83 (m, 1H), 3.42-3.57 (m, 2H),3.28-3.41 (m, 3H), 2.98 (br d, J=14.03 Hz, 1H), 2.60-2.87 (m, 1H),2.17-2.39 (m, 2H), 1.86-1.98 (m, 1H), 1.24-1.52 (m, 2H).

Example 66 Synthesis of(9S)-3-chloro-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(300 mg, 0.997 mmol) in THF (30 mL) under nitrogen at RT was addedtriethylamine (0.834 mL, 5.98 mmol), triphosgene (296 mg, 0.997 mmol)and stirred for 4 h. then pyridin-3-amine (282 mg, 2.99 mmol) was addedand the reaction was heated at 65° C. for 16 h. (TLC eluent: 100% EtOAc:R_(f)-0.3; UV active). The reaction mixture was cooled to RT,concentrated and residue was partitioned between water (5 mL) and EtOAc(20 mL). Organic layer was separated, dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to give crude compound. The crudewas purified by column chromatography (neutral alumina, eluent: 60%ethyl acetate in hexane) to afford the desired product(9S)-3-chloro-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(135 mg, 0.319 mmol, 31.9% yield) as an off-white solid. LCMS (m/z):550.13 [M+H]⁺, R_(t)=1.95 min

¹H NMR (400 MHz, CDCl₃): δ ppm 13.11 (s, 1H), 8.71 (d, J=5.04 Hz, 1H),8.49 (d, J=2.19 Hz, 1H), 8.29 (dd, J=4.60, 1.32 Hz, 1H), 8.05-7.97 (m,1H), 7.59 (s, 1H), 7.54 (s, 1H), 7.51 (d, J=5.04 Hz, 1H), 7.22 (dd,J=8.33, 4.82 Hz, 1H), 5.00 (br s, 1H), 3.42-3.32 (m, 3H), 2.98 (br d,J=14.03 Hz, 1H), 2.70 (s, 3H), 2.25 (br d, J=14.25 Hz, 1H), 2.00-1.87(m, 1H), 1.51-1.40 (m, 2H)

Example 67 Synthesis of(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.331 mmol) in methanol (10 mL) at 0° C. was added aq. HCl(0.201 mL, 6.61 mmol, 36%) and stirred for 2 h. (TLC eluent: 100% EtOAc:R_(f)-0.2; UV active). The reaction mixture was basified with saturatedsodium bicarbonate solution (till pH-8-9) and solvent was evaporatedunder reduced pressure. The residue was diluted with water (10 mL) andextracted into dichloromethane (2×15 mL). Combined organic extracts weredried over anhydrous sodium sulphate, filtered and filtrate wasevaporated in vacuo and the crude was triturated with pentane (10 mL) toafford the desired product(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(123 mg, 0.214 mmol, 64.6% yield) as an off-white solid. LCMS (m/z):565.14 [M+H]⁺, R_(t)=2.33 min

¹H NMR (400 MHz, CDCl₃): δ ppm 13.27 (s, 1H), 8.88 (s, 1H), 8.03 (s,1H), 7.96 (br d, J=7.67 Hz, 1H), 7.92 (s, 1H), 7.78-7.74 (m, 1H),7.72-7.66 (m, 1H), 7.61 (s, 1H), 5.01 (br s, 1H), 3.99-3.92 (m, 1H),3.91-3.83 (m, 2H), 3.66-3.59 (m, 2H), 3.42-3.31 (m, 3H), 2.99 (br d,J=13.59 Hz, 1H), 2.56 (d, J=4.82 Hz, 1H), 2.26 (br d, J=15.35 Hz, 1H),2.07 (br t, J=6.03 Hz, 1H), 2.00-1.86 (m, 1H), 1.52-1.39 (m, 2H)

Example 68 Synthesis of(9S)-3-methyl-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-methyl-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(250 mg, 0.892 mmol) in THF (25 mL) under nitrogen atmosphere at RT wasadded phenyl pyridin-3-ylcarbamate (573 mg, 2.68 mmol), DMAP (327 mg,2.68 mmol) and stirred at 65° C. for 48 h. (TLC eluent: 5% MeOH in DCM:R_(f)-0.4; UV active). The reaction mixture was cooled to RT,concentrated and the residue was partitioned between water (10 mL) andEtOAc (30 mL). Organic layer was separated, dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to give crude compound. The crudewas purified by column chromatography (neutral alumina, eluent: 70%EtOAc in Hexane) to afford(9S)-3-methyl-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(178 mg, 0.437 mmol, 49.0% yield) as a pale yellow solid. LCMS (m/z):401.1 [M+H]⁺, R_(t)=3.342 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.57 (s, 1H), 8.68 (d, J=5.26 Hz, 1H),8.48 (d, J=2.41 Hz, 1H), 8.26 (dd, J=4.71, 1.43 Hz, 1H), 8.08-7.99 (m,1H), 7.42-7.36 (m, 2H), 7.30 (dd, J=5.15, 1.21 Hz, 1H), 7.20 (dd,J=8.33, 4.60 Hz, 1H), 4.97 (br s, 1H), 3.42-3.28 (m, 3H), 3.00 (br d,J=13.59 Hz, 1H), 2.69 (s, 3H), 2.36 (s, 3H), 2.30-2.20 (m, 1H),2.01-1.87 (m, 1H), 1.54-1.35 (m, 2H)

Example 69 Synthesis of(4S)-8-chloro-N-(2-(((R)-2,3-dihydroxypropoxy)pyrimidin-5-yl)-7-(3-(trifluoromethyl)phenyl)-3,4-dihydro-1,4-methanopyrido[2,3-b][1,4]diazepine-5(2H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(150 mg, 0.248 mmol) in methanol (10 mL) at RT was added aq. HCl (1.240mL, 2.479 mmol, 36%) and stirred for 2 h. (TLC eluent: 100% EtOAc:R_(f)-0.2; UV active). The reaction mixture was basified with saturatedsodium bicarbonate solution (till pH 8-9) and solvent was evaporatedunder reduced pressure. The residue was diluted with water (5 mL) andextracted into dichloromethane (2×10 mL). Combined organic extracts weredried over anhydrous sodium sulphate, filtered and filtrate wasevaporated in vacuo and the crude was triturated with diethylether (10mL) and pentane (10 mL) to afford the desired product(9S)-3-chloro-N-(2-((R)-2,3-dihydroxypropoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(89 mg, 0.153 mmol, 61.6% yield) as an off-White solid. LCMS (m/z):565.17 [M+H]⁺, R_(t)=2.30 min

¹H NMR (400 MHz, CDCl₃): δ ppm 13.10 (s, 1H), 8.58 (s, 2H), 8.02 (s,1H), 7.94 (d, J=7.89 Hz, 1H), 7.77 (d, J=7.89 Hz, 1H), 7.71-7.65 (m,1H), 7.61 (s, 1H), 4.98 (br s, 1H), 4.51-4.41 (m, 2H), 4.15-4.06 (m,1H), 3.82-3.67 (m, 2H), 3.44-3.31 (m, 3H), 3.14 (d, J=5.26 Hz, 1H), 2.98(br d, J=13.59 Hz, 1H), 2.34-2.18 (m, 2H), 2.00-1.88 (m, 1H), 1.54-1.42(m, 2H)

Example 70 Synthesis of(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl) methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl) phenyl)-8,9-dihydro-6H-5,9methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (0.25 g, 0.413mmol) in methanol (10 mL) at 0° C. was added aq. HCl (0.517 mL, 6.20mmol, 36%) and stirred at RT for 1 h. (TLC eluent: 100% EtOAc:R_(f)-0.2; UV active). Reaction mixture was basified by adding saturatedsodium bicarbonate solution (till pH-8-9) then concentrated. The residuewas diluted water (10 mL) and extracted into EtOAc (2×25 mL). Combinedorganic extracts were dried over anhydrous Na₂SO₄, filtered and filtratewas evaporated to give crude product. The crude was triturated withdiethyl ether (10 mL) to afford desired product(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.18 g, 0.311 mmol, 75% yield) as off-white solid. LCMS (m/z): 565.14[M+H]⁺, R_(t)=2.51 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.64 (s, 1H), 8.35 (s, 1H), 8.19 (s,1H), 8.11 (br d, J=7.89 Hz, 1H), 7.75 (br d, J=8.11 Hz, 1H), 7.69-7.60(m, 2H), 7.55 (s, 1H), 4.95 (br s, 1H), 4.51-4.46 (m, 2H), 4.06-3.98 (m,1H), 3.75-3.63 (m, 2H), 3.48-3.31 (m, 4H), 2.99 (br d, J=13.59 Hz, 1H),2.51 (br t, J=6.14 Hz, 1H), 2.23 (br d, J=12.28 Hz, 1H), 1.98-1.88 (m,1H), 1.49-1.40 (m, 2H).

Example 71 Synthesis of(9S)-3-chloro-N-cyclopropyl-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of (9S)-phenyl3-chloro-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxylate(400 mg, 0.844 mmol) in THF (30 mL) at RT was added DMAP (516 mg, 4.22mmol), cyclopropanamine (0.301 mL, 4.22 mmol) and stirred at 65° C. for16 h. (TLC eluent: 70% Ethyl acetate in pet ether, Rf: 0.4). Thereaction mixture was cooled to RT, concentrated in vacuo and the residuewas partitioned between water (40 mL) and EtOAc (80 mL). Organic layerwas separated and dried over anhydrous Na₂SO₄, filtered and filtrate wasevaporated to get crude compound. The crude compound was purified bycolumn chromatography (using silica gel, eluent 50% Ethylacetate inhexane) followed by preparative HPLC (Column: XBridge C 18(150×4.6 mm,3.5p), Mobile Phase: A: 0.01 M Ammonium Bicarbonate B: ACN, Gradient:Time/% B: 0/5, 0.8/5, 5/50, 8/95, 12/95, 12.5/5, 15/5, Temp: Ambient,Flow Rate: 0.8 ml/min, Diluent: ACN) to afford(9S)-3-chloro-N-cyclopropyl-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(118 mg, 0.269 mmol, 31.9% yield) as an off white solid. LCMS (m/z):437.05 [M+H]⁺; Rt=2.03 min

¹H NMR (400 MHz, CDCl₃): δ ppm 10.51 (br s, 1H), 7.97 (s, 1H), 7.92 (d,J=7.67 Hz, 1H), 7.71 (d, J=7.67 Hz, 1H), 7.64-7.59 (m, 1H), 7.50 (s,1H), 4.95 (t, J=2.19 Hz, 1H), 3.35-3.26 (m, 3H), 2.89 (br d, J=13.59 Hz,1H), 2.79 (tq, J=7.19, 3.68 Hz, 1H), 2.24-2.16 (m, 1H), 1.92-1.81 (m,1H), 1.52-1.33 (m, 2H), 0.75-0.67 (m, 2H), 0.46-0.40 (m, 2H).

Example 72 Synthesis of(9S)-3-chloro-N-(2-((S)-2,3-dihydroxypropoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (300 mg, 0.496mmol) in methanol (20 mL) at 0° C. was added aq. HCl (1.2 mL, 14.40mmol) and stirred for 4 h. (TLC eluent: 100% EtOAc, Rf: 0.2). Thereaction mixture was concentrated in vacuo and the residue was basifiedwith saturated NaHCO₃ solution (20 mL). The resultant solid was filteredand purified by chiral. (Column/dimensions:Chiralpak AD-H (250×30) mm,5μ % CO2: 50.0% % Co solvent: 50.0% (0.5% DEA in MeOH), Total Flow: 70.0g/min, Back Pressure: 100.0 bar, UV: 260 nm, Stack time: 14. min,Load/inj: 38.0 mg, Solubility: Methanol, Total No of injections: 8Instrument details: Make/Model: Thar SFC-80) to afford(9S)-3-chloro-N-(2-((S)-2,3-dihydroxypropoxy)pyrimidin-5-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(139 mg, 0.239 mmol, 48.1% yield) as an off white solid. LCMS (m/z):565.17 [M+H]⁺, Rt=2.30 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.10 (s, 1H), 8.58 (s, 2H), 8.08-7.91(m, 2H), 7.78 (s, 1H), 7.71-7.57 (m, 2H), 4.98 (br s, 1H), 4.59-4.38 (m,2H), 4.17-4.05 (m, 1H), 3.87-3.63 (m, 2H), 3.47-3.26 (m, 3H), 2.98 (d,J=13.81 Hz, 1H), 2.23 (d, J=14.25 Hz, 2H), 2.04-1.83 (m, 2H), 1.69-1.38(m, 2H)

Example 73 Synthesis of(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.331 mmol) in Methanol (2 mL) was added hydrochloric acid (0.5mL, 16.46 mmol) drop wise over a period of 5 min at 0° C. Then thereaction mixture was stirred at room temperature for 2 h. (TLC eluent:10% MeOH in DCM: R_(f)-0.3). Evaporated the solvent and neutralized withsodium bicarbonate solution filtered the obtained solid and washed withwater to afford the desired product(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(130 mg, 0.229 mmol, 69.2% yield) as a white solid. LCMS (m/z): 565.17[M+H]⁺, Rt=2.51 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.35 (s, 1H), 8.83 (d, J=1.32 Hz, 1H),8.18-8.12 (m, 2H), 7.91-7.85 (m, 3H), 7.83-7.76 (m, 1H), 4.92 (d, J=5.26Hz, 1H), 4.94-4.90 (m, 1H), 4.83 (brs, 1H), 4.63 (t, J=5.70 Hz, 1H),4.30 (dd, J=10.85, 4.06 Hz, 1H), 4.15 (dd, J=10.74, 6.58 Hz, 1H),3.85-3.76 (m, 1H), 3.48-3.37 (m, 3H), 3.32 (d, J=3.07 Hz, 1H), 2.91 (d,J=14.03 Hz, 1H), 2.11-1.76 (m, 2H), 1.50-1.27 (m, 2H).

Example 74 Synthesis of(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (250 mg, 0.414 mmol) inMethanol (8 mL) and Tetrahydrofuran (5 mL) was added HCl (0.5 mL, 16.46mmol) at 0° C. then stirred at RT for 2 h. (TLC eluent: neat ethylacetate, R_(f): 0.2). The reaction mixture was concentrated in vacuo andthe residue was neutralized with saturated NaHCO₃ solution and filteredthe obtained solid, washed with n-pentane (10 mL×3) to afford thedesired product(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(201 mg, 0.354 mmol, 86% yield) as an off white solid. LCMS (m/z):564.17 [M+H]⁺, Rt=2.44 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.34 (s, 1H), 8.22 (s, 1H), 8.14 (d,J=7.89 Hz, 1H), 8.04 (d, J=8.99 Hz, 1H), 7.95 (d, J=3.07 Hz, 1H), 7.73(d, J=7.89 Hz, 1H), 7.67-7.61 (m, 1H), 7.58 (s, 1H), 7.25-7.22 (m, 1H),4.98 (br s, 1H), 4.14-4.03 (m, 3H), 3.89-3.82 (m, 1H), 3.79-3.72 (m,1H), 3.40-3.31 (m, 3H), 2.98 (d, J=13.81 Hz, 1H), 2.54 (d, J=4.38 Hz,1H), 2.25 (d, J=12.06 Hz, 1H), 1.97-1.87 (m, 2H), 1.52-1.35 (m, 2H).

Example 75 Synthesis of(9S)-3-chloro-N-(4-(((S)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(4-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.331 mmol) in methanol (5 mL) and Tetrahydrofuran (5 mL) wasadded HCl (0.3 mL, 9.87 mmol) at 0° C. then stirred at RT for 1 h. (TLCsystem: neat ethyl acetate, R_(f): 0.1). The reaction mixture wasconcentrated in vacuo and the residue was neutralized with saturatedNaHCO₃ solution and filtered the obtained solid, washed with n-pentane(10 mL×2) to afford the desired product(9S)-3-chloro-N-(4-(((S)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(114 mg, 0.200 mmol, 60.4% yield) as an off white solid. LCMS (m/z):565.17 [M+H]⁺, Rt=2.05 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.90 (s, 1H), 8.21-8.16 (m, 2H), 8.10(d, J=7.67 Hz, 1H), 7.73 (d, J=7.89 Hz, 1H), 7.66-7.59 (m, 2H), 6.42 (d,J=5.70 Hz, 1H), 5.03 (br s, 1H), 4.64 (dd, J=11.95, 5.37 Hz, 1H), 4.46(dd, J=11.95, 4.49 Hz, 1H), 4.11 (d, J=6.36 Hz, 1H), 3.94-3.88 (m, 1H),3.64-3.57 (m, 2H), 3.46-3.30 (m, 4H), 2.97 (d, J=14.03 Hz, 1H), 2.27 (d,J=12.50 Hz, 1H), 1.97-1.86 (m, 1H), 1.52-1.39 (m, 2H).

Example 76 Synthesis of(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(240 mg, 0.397 mmol) in Methanol (10 mL) was added aq.HCl (0.345 mL,3.97 mmol) drop wise over a period of 2 min at 0° C. and stirred at roomtemperature for 1 h. (TLC system: 100% Ethyl acetae. Rf value: 0.3).Then the reaction mixture was partitioned between saturated NaHCO₃solution (20 mL) and DCM (30 mL), the separated organic layer was driedover anhydrous sodium sulphate and concentrated under reduced pressureto obtain crude compound. The crude product was purified by flash columnchromatography (100-200 silicagel eluted with 0-2% of MeOH in Ethylacetate) to afford the desired product(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(170 mg, 0.299 mmol, 75% yield) as an off white solid. LCMS (m/z):564.20 [M+H]⁺, Rt=2.17 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.21 (s, 1H), 8.04 (s, 1H), 7.99 (d,J=2.41 Hz, 1H), 7.94 (d, J=7.45 Hz, 1H), 7.88 (t, J=2.30 Hz, 1H), 7.83(d, J=1.97 Hz, 1H), 7.76-7.80 (m, 1H), 7.67-7.74 (m, 1H), 7.61 (s, 1H),4.97 (brs, 1H), 4.01-4.13 (m, 3H), 3.79-3.86 (m, 2H), 3.71-3.77 (m, 1H),3.30-3.42 (m, 3H), 2.99 (br d, J=13.81 Hz, 1H), 2.63 (brs, 1H), 2.24(brd, J=14.69 Hz, 1H), 1.88-1.99 (m, 1H), 1.39-1.48 (m, 1H), 1.26 (s,1H).

Example 77 Synthesis of(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.331 mmol) in Methanol (10 mL) was added aq.HCl (0.287 mL,3.31 mmol), drop wise over a period of 2 min at 0° C. and stirred atroom temperature for 1 h. (TLC system: 100% Ethyl acetae. Rf value:0.4). Then the reaction mixture was partitioned between saturated NaHCO₃solution (20 mL) and DCM (30 mL) dried over anhydrous sodium sulphateand concentrated under reduced pressure to afford the desired product(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyrazin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(171 mg, 0.299 mmol, 91% yield) as an off white solid. LCMS (m/z):565.17 [M+H]⁺, Rt=2.51 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.35 (s, 1H), 8.83 (s, 1H), 8.15 (s,2H), 7.26-7.96 (m, 4H), 4.92 (d, J=5.26 Hz, 1H), 4.83 (br s, 1H), 4.63(br t, J=5.81 Hz, 1H), 4.30 (br dd, J=10.85, 3.84 Hz, 1H), 4.15 (br dd,J=10.63, 6.69 Hz, 1H), 3.81 (br dd, J=10.19, 5.15 Hz, 1H), 3.30-3.63 (m,5H), 2.91 (br d, J=13.37 Hz, 1H), 1.88-2.11 (m, 2H), 1.34 (br d, J=4.38Hz, 2H).

Example 78 Synthesis of(9S)-3-chloro-N-(4-((R)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-3-chloro-N-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(290 mg, 0.479 mmol) in Methanol (10 mL) was added aq.HCl (0.416 mL,4.79 mmol) drop wise over a period of 5 min at 0° C. and stirred at roomtemperature for 1 h. (TLC system: 100% Ethyl acetae. Rf value: 0.3).Then the reaction mixture was partitioned between saturated aq NaHCO₃solution (20 mL) and DCM (30 mL). The separated organic layer was driedover anhydrous sodium sulphate and concentrated under reduced pressureto afford the desired product(9S)-3-chloro-N-(4-((R)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(100 mg, 0.173 mmol, 36.1% yield) as an off white solid. LCMS (m/z):565.10 [M+H]⁺, Rt=2.07 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.42 (s, 1H), 8.31-8.18 (m, 1H),8.17-7.97 (m, 2H), 7.95-7.80 (m, 1H), 6.58-6.41 (m, 1H), 4.92 (br d,J=5.04 Hz, 1H), 4.80 (br s, 1H), 4.62 (br t, J=5.70 Hz, 1H), 4.24-4.13(m, 2H), 4.08 (br dd, J=10.85, 5.81 Hz, 1H), 3.79-3.67 (m, 2H),3.52-3.31 (m, 3H), 3.34-3.30 (m, 2H), 2.90 (br d, J=12.72 Hz, 1H),2.04-1.78 (m, 2H), 1.42-1.19 (m, 2H).

Example 79 Synthesis of(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.497 mmol) in Methanol (10 mL) was added HCl (0.151 mL, 4.97mmol) at 0° C. and stirred at 25° C. for 2 h. (TLC system: 100%ethylacetate, Rf value: 0.2). The reaction mixture was quenched withsaturated NaHCO₃ solution (10 mL) and extracted with DCM (2×30 mL). Thecombined organic layer was dried over anhydrous Na₂SO₄, filtered andfiltrate was evaporated to obtain crude compound. The crude compound wastriturated with n-pentane (3×10 mL). to afford the desired product(9S)-3-chloro-N-(6-((R)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(140 mg, 0.248 mmol, 49.8% yield) as a white solid. LCMS (m/z): 564.17[M+H]⁺, R_(t)=2.42 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 12.92 (s, 1H), 8.09 (d, J=2.85 Hz, 1H),8.03 (s, 1H), 7.96 (d, J=7.67 Hz, 1H), 7.77 (dd, J=8.88, 2.74 Hz, 2H),7.71-7.64 (m, 1H), 7.59 (s, 1H), 6.73 (d, J=8.99 Hz, 1H), 4.98 (brs,1H), 4.43-4.39 (m, 2H), 4.01-3.95 (m, 2H), 3.70-3.61 (m, 2H), 3.42-3.30(m, 3H), 2.98 (d, J=13.81 Hz, 1H), 2.69 (t, J=6.47 Hz, 1H), 2.24 (d,J=12.06 Hz, 1H), 1.98-1.87 (m, 1H), 1.53-1.40 (m, 2H).

Example 80 Synthesis of(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (200 mg, 0.331 mmol) inMethanol (5 mL) was added aq HCl (1.0 mL, 32.9 mmol), over a period of 5min. at 0° C. Then the reaction mixture was stirred at room temperaturefor 1 h. (TLC eluent: 10% MeOH in DCM Rf: 0.3; UV active) and thereaction mixture was poured in to ice cold water (10 mL) the P^(H) ofthe reaction mixture was adjusted to neutral with saturated NaHCO₃solution and extracted with EtoAc (2×25 mL). The combined organic layerwas washed with water (10 mL), brine solution (10 mL) and dried overanhydrous Na₂SO₄, filtered and concentrated under reduced pressure toobtain crude compound. The crude compound was purified by flash columnchromatography (silicagel: 100-200 mesh, Eluent: 5% MeOH in DCM) toafford the desired product(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(110 mg, 0.190 mmol, 57.4% yield) as an off-white solid. LCMS (m/z):564.38 [M+H]⁺, R_(t)=2.17 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.07 (s, 1H), 8.18-8.10 (m, 2H),7.95-7.81 (m, 5H), 7.64 (br s, 1H), 4.96 (d, J=5.04 Hz, 1H), 4.82 (br s,1H), 4.67 (t, J=5.59 Hz, 1H), 3.99 (dd, J=9.32, 3.84 Hz, 1H), 3.87-3.75(m, 2H), 3.45 (t, J=5.59 Hz, 3H), 3.40 (s, 1H), 2.89 (d, J=13.59 Hz,1H), 2.04-1.90 (m, 2H), 1.41-1.29 (m, 3H).

Example 81 Synthesis of(9S)-3-chloro-N-(2-((S)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a solution of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(170 mg, 0.281 mmol) in methanol (10 mL) under nitrogen at 0° C. wasadded aq. HCl (1 mL, 32.9 mmol, 36%) and stirred at RT for 2 h. (TLCeluent: 100% Ethyl acetate: R_(f)-0.2; UV active). The reaction mixturewas basified with saturated sodium bicarbonate solution (till pH-8-9) at0° C. and solvent was evaporated under reduced pressure. The residue wasdiluted with water (30 mL) and extracted into DCM (2×100 mL). Combinedorganic extracts were dried over anhydrous sodium sulphate, filtered andfiltrate was evaporated to give crude as a white solid. The crude wastriturated with ether and filtered to afford(9S)-3-chloro-N-(2-((S)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(70 mg, 0.121 mmol, 43.0% yield) as off white solid. LCMS (m/z): 565.17[M+H]⁺, R_(t)=2.32 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.57 (s, 1H), 8.32 (d, J=5.70 Hz, 1H),8.15-7.97 (m, 2H), 7.87-7.67 (m, 3H), 7.62 (s, 1H), 4.95 (br s, 1H),4.31-4.07 (m, 2H), 4.00-3.85 (m, 1H), 3.77-3.52 (m, 2H), 3.43-3.21 (m,3H), 3.16 (br d, J=5.26 Hz, 1H), 3.00 (br d, J=13.81 Hz, 1H), 2.41 (brt, J=6.25 Hz, 1H), 2.24 (br d, J=14.69 Hz, 1H), 2.03-1.85 (m, 1H),1.54-1.37 (m, 2H).

Example 82 Synthesis of(9S)-3-chloro-N-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (180 mg, 0.298 mmol) inMethanol (10 mL) was added hydrochloric acid (5 mL, 165 mmol) at 0° C.over a period of 5 min. Then the reaction mixture was stirred at 30° C.for 2 h. (TLC eluent: 5% MeOH in DCM: R_(f)-0.5; UV active). The solventwas evaporated and the reaction mixture was neutralized with sodiumbicarbonate solution, filtered the obtained solid and washed with water(10 mL) and with n-pentane (2×20 mL) to afford the desired product(9S)-3-chloro-N-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(120 mg, 0.212 mmol, 35.5% yield) as an off-white solid. LCMS (m/z): 564[M+H]⁺, Rt=6.0 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.43 (s, 1H), 8.23 (s, 1H), 8.16 (d,J=8.11 Hz, 1H), 8.04 (d, J=5.70 Hz, 1H), 7.71-7.79 (m, 2H), 7.60-7.67(m, 1H), 7.58 (s, 1H), 6.53 (dd, J=5.70, 2.19 Hz, 1H), 4.96 (s, 1H),4.08-4.18 (m, 2H), 3.79-3.87 (m, 1H), 3.70-3.77 (m, 1H), 3.31-3.40 (m,3H), 3.00 (d, J=13.59 Hz, 1H), 2.60 (s, 3H), 2.24 (d, J=12.93 Hz, 1H),2.05 (d, J=7.67 Hz, 1H), 1.86-1.98 (m, 1H), 1.38-1.52 (m, 1H).

Example 83 Synthesis of(9S)-3-chloro-N-(2-((S)-2,3-dihydroxypropoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(2-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide (420 mg, 0.695 mmol) inMethanol (10 mL) was added hydrochloric acid (5 mL, 165 mmol) at 0° C.over a period of 5 min. Then the reaction mixture was stirred at 30° C.for 30 min. (TLC System: 5% MeOH in DCM: R_(f)-0.5; UV active). Then thesolvent was evaporated and the reaction mixture was neutralized withsodium bicarbonate solution, filtered and washed with water andn-pentane (2×20 mL) to afford the desired product(9S)-3-chloro-N-(2-((S)-2,3-dihydroxypropoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(299 mg, 0.517 mmol, 37.1% yield) as an off-white solid. LCMS (m/z):564.20 [M+H]⁺, Rt=2.35 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.29 (s, 1H), 8.02 (s, 1H), 7.97 (d,J=7.67 Hz, 1H), 7.85 (d, J=5.70 Hz, 1H), 7.81 (d, J=7.45 Hz, 1H),7.73-7.65 (m, 1H), 7.61 (s, 1H), 7.06 (d, J=1.53 Hz, 1H), 6.69 (dd,J=5.70, 1.75 Hz, 1H), 4.97 (s, 1H), 4.46-4.42 (m, 2H), 4.28 (d, J=4.82Hz, 1H), 3.96 (d, J=4.38 Hz, 1H), 3.69-3.60 (m, 2H), 3.41-3.32 (m, 3H),2.98 (d, J=13.81 Hz, 1H), 2.85 (t, J=6.36 Hz, 1H), 2.23 (d, J=14.03 Hz,1H), 2.01-1.87 (m, 1H), 1.49-1.38 (m, 2H).

Example 84 Synthesis of(9S)-3-chloro-N-(6-(((S)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(150 mg, 0.248 mmol) in Methanol (10 mL) was added Hydrochloric acid (4mL, 132 mmol) over a period of 5 min. at 0° C. Then the reaction mixturewas stirred at 30° C. for 30 min. (TLC eluent: 5% MeOH in DCM:R_(f)-0.5;

UV active). The solvent was evaporated and reaction mixture wasneutralized with sodium bicarbonate solution, extracted with DCM (2×50.mL). The combined organic layer washed with brine solution and driedover anhydrous sodium sulphate, filtered and evaporated to obtain crudecompound. The crude product was purified by combi-flash chromatography(120 g reverse phase column: Eluent: 100% acetonitrile) and the obtainedproduct was washed with n-pentane (2×20 mL) to afford the desiredproduct(9S)-3-chloro-N-(6-((S)-2,3-dihydroxypropoxy)pyridin-3-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(108 mg, 0.190 mmol, 77% yield) as an off white solid. LC-MS (m/z):564.17 [M+H]⁺, Rt=2.42 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 12.92 (s, 1H), 8.09 (d, J=2.85 Hz, 1H),8.03 (s, 1H), 7.96 (d, J=7.67 Hz, 1H), 7.77 (dd, J=8.99, 2.63 Hz, 2H),7.71-7.63 (m, 1H), 7.59 (s, 1H), 6.73 (d, J=8.77 Hz, 1H), 4.99 (s, 1H),4.42 (d, J=4.17 Hz, 2H), 3.97 (s, 2H), 3.73-3.61 (m, 2H), 3.40-3.31 (m,3H), 2.98 (d, J=13.59 Hz, 1H), 2.67 (t, J=6.14 Hz, 1H), 2.23 (d, J=14.03Hz, 1H), 1.99-1.87 (m, 1H), 1.49-1.38 (m, 2H).

Example 85 Synthesis of(9S)-3-chloro-N-(2-(((R)-2,3-dihydroxypropoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred suspension of(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.414 mmol) in methanol (5.0 mL) was added aq. HCl (0.349 mL,4.14 mmol, 36%) at 0° C. and stirred at RT for 5 h. After completion ofthe reaction the volatiles were evaporated under reduced pressure to getthe crude (TLC eluent system: 5% Methanol in DCM, Rf-0.2, UV active).The crude was diluted with water (5 ml) and basified (up to pH 8) withthe aqueous sodium bicarbonate solution. The precipitated solid wasfiltered, washed with the water and dried under vacuum to afford(9S)-3-chloro-N-(2-((R)-2,3-dihydroxypropoxy)pyridin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(149.0 mg, 0.262 mmol, 63.4% yield) as an off white solid. LCMS (m/z):564.20 [M+H]⁺, R_(t)=2.37 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.30 (s, 1H), 8.02 (s, 1H), 7.97 (br d,J=7.67 Hz, 1H) 7.78-7.88 (m, 2H), 7.66-7.73 (m, 1H), 7.61 (s, 1H), 7.06(d, J=1.32 Hz, 1H), 6.69 (dd, J=5.70, 1.53 Hz, 1H), 4.96 (br s, 1H),4.43 (d, J=4.60 Hz, 2H), 4.30 (br s, 1H), 3.97 (br d, J=3.73 Hz, 1H),3.65 (br d, J=4.38 Hz, 2H), 3.29-3.42 (m, 3H), 2.87-3.01 (m, 2H), 2.23(br d, J=14.47 Hz, 1H), 1.87-2.00 (m, 1H), 1.39-1.55 (m, 2H).

Example 86 Synthesis of(9S)-3-chloro-N-(6-(((S)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(6-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(200 mg, 0.331 mmol) in Methanol (5 mL) at 0° C. was added hydrochloricacid (1 mL, 32.9 mmol) drop wise over a period of 5 min. Then thereaction mixture was stirred at 28° C. for 30 min. (TLC eluent: 10% MeOHin DCM: R_(f)-0.2; UV active). The reaction mixture was neutralized withsodium bicarbonate solution and filtered the obtain solid, trituratedwith diethylether to afford the desired product(9S)-3-chloro-N-(6-(((S)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(110 mg, 0.189 mmol, 57.1% yield) as an off-white solid. LCMS (m/z):565.10 [M+H]⁺, R_(t): 2.52 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.64 (s, 1H), 8.35 (d, J=0.66 Hz, 1H),8.19 (s, 1H), 8.11 (d, J=7.89 Hz, 1H), 7.75 (d, J=8.11 Hz, 1H),7.67-7.62 (m, 1H), 7.61 (s, 1H), 7.55 (d, J=0.88 Hz, 1H), 4.95 (br s,1H), 4.55-4.42 (m, 2H), 3.98-4.08 (m, 1H), 3.75-3.62 (m, 2H), 3.50-3.31(m, 4H), 2.99 (d, J=13.59 Hz, 1H), 2.52 (t, J=6.25 Hz, 1H), 2.22 (br d,J=14.69 Hz, 1H), 2.02-1.82 (m, 1H), 1.53-1.37 (m, 2H).

Example 87 Synthesis of(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.414 mmol) in Methanol (5 mL) was added aq. HCl (0.126 mL,4.14 mmol) at 0° C. The reaction mixture was stirred for 1 h. (TLCSystem: R_(f): 0.4, EtOAc). at room temperature and concentrated underreduced pressure to obtain residue. The residue was neutralized withsaturated sodium bicarbonate solution and filtered the obtained solidand dried to afford the desired product(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(160 mg, 0.274 mmol, 66.1% yield) as an off white solid. LCMS (m/z):564.1 [M+H]⁺, R_(t)=2.44 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.22 (s, 1H), 8.23-8.12 (m, 2H), 7.99(d, J=9.21 Hz, 1H), 7.90-7.84 (m, 3H), 7.83-7.78 (m, 1H), 7.43 (dd,J=9.21, 3.07 Hz, 1H), 4.95 (br s, 1H), 4.83 (br s, 1H), 4.65 (br s, 1H),4.02 (dd, J=9.87, 3.95 Hz, 1H), 3.92-3.85 (m, 1H), 3.82-3.74 (m, 1H),3.48-3.31 (m, 5H), 2.89 (d, J=13.81 Hz, 1H), 2.06-1.89 (m, 2H),1.42-1.30 (m, 2H).

Example 88 Synthesis of(9S)-3-chloro-N-(2-(((R)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(2-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.496 mmol) in methanol (10 mL) under nitrogen at 0° C. wasadded aq. HCl (1 ml, 4.00 mmol, 36%) and stirred at RT for 1 h. (TLCeluent: 5% Methanol in DCM, R_(f): 0.3, UV active). To the reactionmixture was added saturated NaHCO₃ solution (till pH-8-9) and extractedinto EtOAc (3×10 mL). The combined organic extracts were dried overanhydrous Na₂SO₄, filtered and filtrate was evaporated to obtain crudeproduct. The crude compound was purified by chromatography (GRACEinstrument using C-18 column, Mobile phase A: 0.1% Formic Acid in water;B: ACN, the product was eluted at 90% ACN/0.1% Formic Acid in water.)Subsequently required fractions were concentrated and basified withsaturated NaHCO₃ and extracted in to DCM. Combined extracts were driedover anhydrous Na₂SO₄, filtered and evaporated to afford(9S)-3-chloro-N-(2-((R)-2,3-dihydroxypropoxy)pyrimidin-4-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(170 mg, 0.292 mmol, 58.9% yield) as an off-white solid. LCMS (m/z):565.2 [M+H]⁺, R_(t)=2.30 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.57 (s, 1H), 8.32 (d, J=5.70 Hz, 1H),8.10-8.05 (m, 2H), 7.79-7.69 (m, 3H), 7.62 (s, 1H), 4.95 (br s, 1H),4.24-4.12 (m, 2H), 3.97 (dq, J=10.06, 5.05 Hz, 1H), 3.72-3.58 (m, 2H),3.41-3.32 (m, 3H), 3.11 (d, J=5.26 Hz, 1H), 3.00 (br d, J=14.03 Hz, 1H),2.37 (t, J=6.36 Hz, 1H), 2.24 (br d, J=14.03 Hz, 1H), 2.00-1.88 (m, 1H),1.53-1.39 (m, 2H)

Example 89 Synthesis of(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(120 mg, 0.198 mmol) in methanol (15 mL) was added aq.HCl (6.03 μL,0.198 mmol, 36%) at 0° C. and stirred at RT for 1 h. (TLC eluent: 10%MeOH in EtOAc: R_(f)-0.1; UV active). The reaction mixture was basifiedwith saturated sodium bicarbonate solution (till pH-8-9) at 0° C. andconcentrated. The residue was diluted with water (8 mL) and extractedinto DCM (2×25 mL). Combined organic extracts were dried over anhydroussodium sulphate, filtered and filtrate was evaporated under reducedpressure to afford(9S)-3-chloro-N-(5-((R)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(40 mg, 0.068 mmol, 34.4% yield) as brown solid. LCMS (m/z): 565.17[M+H]⁺, R_(t)=2.25 min.

¹H NMR (400 MHz, DMSO-d₆): δ ppm 13.43 (s, 1H), 8.33 (s, 2H), 8.00-8.24(m, 2H), 7.83-7.91 (m, 2H), 7.83-7.91 (m, 1H), 5.01 (br d, J=3.95 Hz,1H), 4.80 (br s, 1H), 4.47-4.74 (m, 1H), 4.12 (dd, J=10.08, 3.73 Hz,2H), 3.89-4.06 (m, 1H), 3.36-3.48 (m, 3H), 3.30-3.34 (m, 2H), 2.89 (brd, J=13.59 Hz, 1H), 1.98 (br s, 2H), 1.35 (br s, 2H).

Example 90 Synthesis of(9S)-4-methyl-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-4-methyl-2-(2-methylpyridin-4-yl)-7,8,9,10-tetrahydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine(500 mg, 1.783 mmol) in THF (30 mL) at RT was added DIPEA (0.934 mL,5.35 mmol), triphosgene (318 mg, 1.070 mmol) and stirred for 1 h. thenadded pyridin-3-amine (252 mg, 2.68 mmol) and the reaction mixture washeated to 75° C. for 16 h. (TLC eluting system: 10% MeOH in DCM;R_(f)-0.3; UV active). The reaction mixture was cooled to RT andquenched with water (15 mL) and extracted into EtOAc (2×15 mL). Organiclayer was separated, dried over anhydrous sodium sulphate, filtered andfiltrate was evaporated to get crude compound. The crude was purified bychromatography (GRACE using C-18 reserval column, Mobile phase A: 0.1%Formic Acid in water; B: ACN, eluent 56% B in A). Combined fractionswere evaporated and basified with saturated NaHCO₃ solution. The aqueouslayer was extracted with DCM, DCM layer was dried over anhydrous Na₂SO₄,filtered and filtrate was evaporated to afford(9S)-4-methyl-2-(2-methylpyridin-4-yl)-N-(pyridin-3-yl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(160 mg, 0.395 mmol, 22.17% yield) as yellow solid. LCMS (m/z): 400.9[M+H]⁺, R_(t)=4.45 min.

¹H NMR (400 MHz, CDCl₃): δ ppm 13.83 (s, 1H), 8.57-8.76 (m, 2H), 8.31(dd, J=4.71, 1.43 Hz, 1H), 8.02-8.23 (m, 1H), 7.61 (s, 1H), 7.51 (dd,J=5.04, 1.53 Hz, 1H), 7.20-7.33 (m, 2H), 4.99 (br s, 1H), 3.38 (dd,J=13.59, 1.75 Hz, 1H), 3.12-3.28 (m, 2H), 2.98 (br d, J=13.59 Hz, 1H),2.68 (s, 3H), 2.44 (s, 3H), 2.13-2.41 (m, 1H), 1.85-2.00 (m, 1H), 1.44(dt, J=6.08, 3.21 Hz, 2H).

Example 91 Synthesis of(9S)-N-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-3-methyl-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-N-(4-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)-3-methyl-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(300 mg, 0.514 mmol) in methanol (10 mL) was added HCl (1 mL, 32.9 mmol)drop wise over a period of 5 min. at 0° C. Then the reaction mixture wasstirred at 30° C. for 1 h. (TLC eluent: 5% MeOH in DCM: R_(f)-0.3; UVactive).

Evaporated the solvent under reduced pressure and neutralized withsodium bicarbonate solution and filtered the obtain solid, it wastriturated with 1:1 ratio of diethylether (50 ml) and pentane (50 ml) toafford pure compound(9S)-N-(4-((R)-2,3-dihydroxypropoxy)pyridin-2-yl)-3-methyl-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(250 mg, 0.453 mmol, 88% yield) as an off white solid. LCMS (m/z):544.26 [M+H]⁺, R_(t)=2.23 min.

¹H NMR (400 MHz, CDCl₃): δ: 13.76 (s, 1H), 8.08-8.00 (m, 2H), 7.97-7.88(m, 2H), 7.73-7.57 (m, 2H), 7.38 (d, J=0.7 Hz, 1H), 7.23 (dd, J=9.0, 3.1Hz, 1H), 4.95 (d, J=3.1 Hz, 1H), 4.16-4.00 (m, 3H), 3.85 (dd, J=11.4,3.8 Hz, 2H), 3.76 (dd, J=11.4, 5.4 Hz, 3H), 3.01 (d, J=13.6 Hz, 1H),2.55-2.4 (m, 1H), 2.39 (s, 3H), 2.25 (d, J=14.4 Hz, 1H), 1.98-1.84 (m,2H), 1.42-1.33 (m, 1H).

Example 92 Synthesis of(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide

To a stirred solution of(9S)-3-chloro-N-(5-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.420 g, 0.694 mmol) in methanol (5 mL) was added aq. HCl (0.5 mL, 6.00mmol, 12M) at 0° C. and stirred at RT for 1 h. (TLC eluent 100%Ethylacetate: R_(f) ⁼0.1; UV active). Reaction mixture was basified byadding saturated sodium bicarbonate solution (till pH-8-9) thenconcentrated. The residue was diluted with water (10 mL) and extractedinto ethyl acetate (20 mL). Combined organic extracts were dried overanhydrous sodium sulphate, filtered and filtrate was evaporated in vacuoand the crude was triturated with diethylether (10 mL) to afford thedesired product(9S)-3-chloro-N-(5-((S)-2,3-dihydroxypropoxy)pyrimidin-2-yl)-2-(3-(trifluoromethyl)phenyl)-8,9-dihydro-6H-5,9-methanopyrido[2,3-b][1,4]diazocine-10(7H)-carboxamide(0.083 g, 0.145 mmol, 20.90% yield) as an off white solid. LCMS (m/z):565.14 [M+H]⁺, R_(t)=2.25 min.

¹H NMR (400 MHz, CDCl₃): δ 13.74 (s, 1H), 8.29 (s, 2H), 8.18 (s, 1H),8.10 (d, J=7.89 Hz, 1H), 7.77-7.75 (m, 1H), 7.65-7.56 (m, 2H), 5.03 (brs, 1H), 4.21-4.00 (m, 3H), 3.86 (br dd, J=11.18, 2.63 Hz, 1H), 3.76 (brdd, J=11.18, 4.82 Hz, 1H), 3.44-3.24 (m, 3H), 2.98 (br d, J=13.59 Hz,1H), 2.62 (br s, 1H), 2.38-2.21 (m, 1H), 2.01 (br s, 2H), 1.98-1.75 (m,2H).

Example 93. Full-Length SIRT1 Production

Full-length human SIRT1 (hSIRT1) proteins were expressed with aC-terminal His₆ tag and purified as described in Hubbard. et al. (2013)Science 339, 1216. Each cell paste was resuspended in buffer A (50 mMTris-HCl pH 7.5, 250 mM NaCl, 25 mM imidazole, and 0.1 mM TCEP) with1,000 U Benzonase nuclease (Sigma Aldrich) supplemented with cOmplete,EDTA-free Protease Inhibitor Cocktail Tablets (Roche) on ice. Cells weredisrupted by pulse sonication with 50% on and 50% off for 12 minutestotal at 40 W. Insoluble debris was removed by centrifugation. Clarifiedsupernatant was directly loaded onto a 1 mL HisTrap FF Crude column (GELifesciences). After washing with buffer A, SIRT1 was eluted with bufferB (50 mM Tris-HCl pH 7.5, 250 mM NaCl, 500 mM imidazole and 0.1 mMTCEP). Protein was further purified by size exclusion chromatography inbuffer C (50 mM Tris-HCl pH 7.5, 300 mM NaCl, 0.1 mM TCEP) using aHi-load Superdex 200 16/60 column (GE Lifesciences). Enzymeconcentrations were determined by Bradford assay using BSA as astandard. Final protein purity was assessed by gel densitometry.Proteins were confirmed by LC/MS. All proteins were greater than 90%pure.

Example 94. SIRT1 Deacetylation Reactions

SIRT1 deacetylation reactions were performed in reaction buffer (50 mMHEPES-NaOH, pH 7.5, 150 mM NaCl, 1 mM DTT, and 1% DMSO) at 25° C.monitoring either nicotinamide production using the continuous PNC1/GDHcoupled assay (Smith, B. C. et al. (2009) Anal Biochem 394, 101) orO-acetyl ADP ribose (OAcADPr) production by mass spectrometry (Hubbard.et al. (2013) Science 339, 1216). Final concentrations of the PNC1/GDHcoupling system components used were 20 units/mL bovine GDH(Sigma-Aldrich), 1 uM yeast PNC1, 3.4 mM α-ketoglutarate, and 220 μMNADH or NADPH. An extinction coefficient of 6.22 mM⁻¹cm⁻¹ and apathlength of 0.81 cm was used to convert the absorbance at 340 nm toproduct concentration for the 150 uL reactions used. Assays monitoringOAcADPr production were performed in reaction buffer with 0.05% BSA andtime points were taken by quenching the deacetylation reaction with astop solution which gave a final concentration of 1% formic acid and 5mM nicotinamide. Quenched reactions were diluted 5-fold with 1:1acetonitrile:methanol and spun at 5,000×g for 10 minutes to precipitateprotein before being analyzed with an Agilent RapidFire 200High-Throughput Mass Spectrometry System (Agilent, Wakefield, Mass.)coupled to an ABSciex API 4000 mass spectrometer fitted with anelectrospray ionization source. The p53-based Ac-p53(W5)(Ac-RHKK^(Ac)W-NH₂) and TAMRA(Ac-EE-K(biotin)-GQSTSSHSK(Ac)NleSTEG-K(5TMR)-EE-NH₂) peptides wereobtained from American Century Peptide and Biopeptide, Inc,respectively.). Substrate K_(M) determinations were performed by varyingone substrate concentration at a fixed, saturating concentration of thesecond substrate. SIRT1 activation and inhibition assays were run inreaction buffer with 0.05 BSA at 25° C. and analyzed using the OAcADPrassay. Enzyme and compound were pre-incubated for 20 minutes beforeaddition of substrates. For the activation screen of full-length hSIRT1,compounds were tested in duplicate with a dose response. In order to besensitive to K_(M)-modulating activators, substrate concentrations ofapproximately one-tenth their K_(M) values were used. Thedose-dependence of five compounds was tested and the fold-activationdata were described by Eq. 1

$\begin{matrix}{\mspace{79mu} {{\frac{v_{x}}{v_{0}} = {b + \frac{{RV}_{{ma}\; x} - b}{1 + \frac{\text{?}}{\text{?}}}}}{\text{?}\text{indicates text missing or illegible when filed}}}} & \left( {{Eq}.\mspace{14mu} 1} \right)\end{matrix}$

where v_(x)/v₀ is the ratio of the reaction rate in the presence (v_(x))versus absence (v₀) of activator (X), RV_(max) is the relative velocityat infinite activator concentration, EC₅₀ is the concentration ofactivator required to produce one-half RV_(max) and b is the minimumvalue of v_(x)/v₀.

Example 95. Biochemical Activity

Mass spectrometry based assays were used to identify modulators of SIRT1activity. The TAMRA based assay utilized a peptide having 20 amino acidresidues as follows: Ac-EE-K(biotin)-GQSTSSHSK(Ac)NleSTEG-K(5TMR)-EE-NH₂(SEQ ID NO: 1), wherein K(Ac) is an acetylated lysine residue and Nle isa norleucine. The peptide was labeled with the fluorophore 5TMR(excitation 540 nm/emission 580 nm) at the C-terminus. The sequence ofthe peptide substrate was based on p53 with several modifications. Inaddition, the methionine residue naturally present in the sequence wasreplaced with the norleucine because the methionine may be susceptibleto oxidation during synthesis and purification. The Trp based assayutilized a peptide having an amino acid residues as follows:Ac-R-H-K-K(Ac)-W-NH2 (SEQ ID NO: 2).

The TAMRA based mass spectrometry assay was conducted as follows: 0.5 μMpeptide substrate and 120 μM βNAD⁺ was incubated with 10 nM SIRT1 for 25minutes at 25° C. in a reaction buffer (50 mM Tris-acetate pH 8, 137 mMNaCl, 2.7 mM KCl, 1 mM MgCl₂, 5 mM DTT, 0.05% BSA). The SIRT1 proteinwas obtained by cloning the SirT1 gene into a T7-promoter containingvector, which was then transformed and expressed in BL21(DE3) bacterialcells. Test compound was added at varying concentrations to thisreaction mixture and the resulting reactions were monitored. After the25 minute incubation with SIRT1, 10 μL of 10% formic acid was added tostop the reaction. The resulting reactions were sealed and frozen forlater mass spec analysis. Determination of the amount of deacetylatedsubstrate peptide formed (or, alternatively, the amount ofO-acetyl-ADP-ribose (OAADPR) generated) by the sirtuin-mediatedNAD-dependent deacetylation reaction allowed for the precise measurementof relative SIRT1 activity in the presence of varying concentrations ofthe test compound versus control reactions lacking the test compound.

The Trp mass spectrometry assay was conducted as follows. 0.5 μM peptidesubstrate and 120 μM βNAD⁺ were incubated with 10 nM SIRT1 for 25minutes at 25° C. in a reaction buffer (50 mM HEPES pH 7.5, 1500 mMNaCl, 1 mM DTT, 0.05% BSA). The SIRT1 protein was obtained by cloningthe SirT1 gene into a T7-promoter containing vector, which was thenexpressed in BL21(DE3) bacterial cells and purified as described infurther detail below. Test compound was added at varying concentrationsto this reaction mixture and the resulting reactions were monitored.After the 25 minute incubation with SIRT1, 10 μL of 10% formic acid wasadded to stop the reaction. The resulting reactions were sealed andfrozen for later mass spec analysis. The relative SIRT1 activity wasthen determined by measuring the amount of O-acetyl-ADP-ribose (OAADPR)formed (or, alternatively, the amount of deacetylated Trp peptidegenerated) by the NAD-dependent sirtuin deacetylation reaction in thepresence of varying concentrations of the test compound versus controlreactions lacking the test compound. The degree to which the test agentactivated deacetylation by SIRT1 was expressed as EC_(1.5) (i.e., theconcentration of compound required to increase SIRT1 activity by 50%over the control lacking test compound), and Percent Maximum Activation(i.e., the maximum activity relative to control (100%) obtained for thetest compound).

A control for inhibition of sirtuin activity was conducted by adding 1μL of 500 mM nicotinamide as a negative control at the start of thereaction (e.g., permits determination of maximum sirtuin inhibition). Acontrol for activation of sirtuin activity was conducted using 10 nM ofsirtuin protein, with 1 μL of DMSO in place of compound, to determinethe amount of deacetylation of the substrate at a given time pointwithin the linear range of the assay. This time point was the same asthat used for test compounds and, within the linear range, the endpointrepresents a change in velocity.

For the above assay, SIRT1 protein was expressed and purified asfollows. The SirT1 gene was cloned into a T7-promoter containing vectorand transformed into BL21(DE3). The protein was expressed by inductionwith 1 mM IPTG as an N-terminal His-tag fusion protein at 18° C.overnight and harvested at 30,000×g. Cells were lysed with lysozyme inlysis buffer (50 mM Tris-HCl, 2 mM Tris[2-carboxyethyl] phosphine(TCEP), 10 μM ZnCl₂, 200 mM NaCl) and further treated with sonicationfor 10 min for complete lysis. The protein was purified over a Ni-NTAcolumn (Amersham) and fractions containing pure protein were pooled,concentrated and run over a sizing column (Sephadex S200 26/60 global).The peak containing soluble protein was collected and run on anIon-exchange column (MonoQ). Gradient elution (200 mM-500 mM NaCl)yielded pure protein. This protein was concentrated and dialyzed againstdialysis buffer (20 mM Tris-HCl, 2 mM TCEP) overnight. The protein wasaliquoted and frozen at −80° C. until further use.

Sirtuin-modulating compounds of Formula (I) that activated SIRT1 wereidentified using the assay described above and are shown below inTable 1. The EC_(1.5) values represent the concentration of testcompounds that result in 150% activation of SIRT1. The EC_(1.5) valuesfor the activating compounds of Formula (I) are represented by A(EC_(1.5)<1 μM), B (EC_(1.5) 1-25 μM), C (EC_(1.5)>25 μM). The percentmaximum fold activation is represented by A (Fold activation≥150%) or B(Fold Activation≤150%). “NT” means not tested; “ND” means notdeterminable. The compound numbering in the table starts with compoundnumber 10, and parenthetic numbering (#) corresponding to the STACnumbering system in FIG. 4 and Examples 90-106 (i.e., compound no. 68 isalso STAC 1, so it is shown as 68(1), and further STACs: 546(3), 444(4),314(5), 816(7), 76(8), and 81(9)).

It is noted that compounds of the present invention have been named bytwo different chemical nomenclature conventions as generated by twodifferent chemical drawing and/or chemical naming computer programs,i.e., generated by Chem Axon (JChem-Excel) and Cambridge Soft(ChemDraw®), respective companies.

TABLE 1 Chemical Name: TRP Example Generated by TRP MAX number StructureCHemAxon Activity RESP 1

(9S)-N-(pyridin-2-yl)- 5-[(2S)-2- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide B A 2

(9S)-N-(5-fluoropyridin-3- yl)-5-[2- (trifluoromethyl)pyridin-4-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 3

(9S)-N-(pyrimidin-5- yl)-5-[5- (trifluoromethyl)pyridin- 3-yl]-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 4

(9S)-5-(2- methylpyridin-4-yl)-N- (pyridin-2-yl)-1,6,8-triazatricyclo[7.3.1.0^(2,7)] trideca-2(7),3,5-triene- 8-carboxamide A A5

(9S)-N-(5- fluoropyridin-3-yl)-5- (2-methylpyridin-4-yl)- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2,4,6-triene-8- carboxamide A A 6

(9S)-N-(pyridazin-3- yl)-5-[2- (trifluoromethyl)pyridin- 4-yl]-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 7

(9S)-N-(pyridin-2-yl)-5-[2- (trifluoromethyl)pyridin-4- yl]-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 8

(9S)-N-(pyridin-2-yl)- 5-[(3S)-3- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide B A 9

(9S)-N-(pyrimidin-4- yl)-5-[2- (trifluoromethyl)pyridin- 4-yl]-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 10

(9S)-N-(pyrimidin-5- yl)-5-[2- (trifluoromethyl)pyridin- 4-yl]-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 11

(9S)-N-(pyridin-2-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 12

(9S)-5-(2- methylpyridin-4-yl)-N- (pyrimidin-4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 13

(9S)-5-(2- methylpyridin-4-yl)-N- (pyrazin-2-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 14

(9S)-5-(2- methylpyridin-4-yl)-N- (pyridazin-3-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 15

(9S)-N-(2-methyl-2H- indazol-5-yl)-5-(2- methylpyridin-4-yl)- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2,4,6-triene-8- carboxamide A A 16

(9S)-5-(2- methylpyridin-4-yl)-N- (pyrimidin-5-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide B A 17

(9S)-N-(pyrazin-2-yl)- 5-[2- (trifluoromethyl)pyridin- 4-yl]-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 18

(9S)-N-(pyrimidin-2- yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 19

(9S)-N-(dimethyl-1,3- thiazol-2-yl)-5-(2- methylpyridin-4-yl)- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 20

(9S)-N-(6- methoxypyrazin-2-yl)- 5-(2-methylpyridin-4- yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 21

(9S)-N-(6-methylpyrazin- 2-yl)-5-(2-methylpyridin- 4-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide B A 22

(9S)-N-(pyrimidin-2- yl)-5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 23

(9S)-N-(pyridin-2-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide B A 24

(9S)-N-(5- fluoropyridin-2-yl)-5- [(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 25

(9S)-N-(pyridin-3-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 26

(9S)-N-(1-methyl-1H- pyrazol-4-yl)-5-(2- methylpyridin-4-yl)- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide B A 27

(9S)-N-(pyrazin-2-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 28

(9S)-N-(pyridin-3-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 29

(9S)-N-(5- fluoropyridin-2-yl)-5- [(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide B A 30

(9S)-N-(pyrazin-2-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide A A 31

(9S)-N-(pyridin-2-yl)- 5-[(3S)-3- (trifluoromethyl)pyrrolidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-8-carboxamide hydrochloride A A 32

(9S)-5-(2- methylpyridin-4-yl)-N- (pyridin-3-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 33

(9S)-4-chloro-5-N- cyclopropyl-8-N- (pyridin-2-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide BA 34

(9S)-8-N-{4-[(2R)-2,3- dihydroxypropoxy] pyridin-2-yl}-5-N-[(2R)-1,1,1-trifluoropropan- 2-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 35

(9S)-5-N-(2,2- difluoropropyl)-8-N- {4-[(2R)-2,3- dihydroxypropoxy]pyridin-2-yl}-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-5,8-dicarboxamide A A 36

(9S)-5-N-cyclopropyl- 8-N-(pyridin-2-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide AA 37

(9S)-8-N-(pyridin-2- yl)-5-N-(2,2,2- trifluoroethyl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide AA 38

(9S)-4-chloro-8-N- (pyridin-2-yl)-5-N- (2,2,2-trifluoroethyl)- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide AA 39

(9S)-4-chloro-8-N- (pyridin-2-yl)-5-N- [(2R)-1,1,1-trifluoropropan-2-yl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 40

(9S)-8-N-(1,2-oxazol- 3-yl)-5-N-[(2R)-1,1,1- trifluoropropan-2-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-5,8-dicarboxamide A A 41

(9S)-8-N-{6-methyl- 1H-pyrazolo[3,4- b]pyridin-3-yl}-5-N-(2,2,2-trifluoroethyl)- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 42

(9S)-5-N-cyclopropyl- 8-N-{6-methyl-1H- pyrazolo[3,4-b]pyridin-3-yl}-1,6,8- triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene-5,8-dicarboxamide A A 43

(9S)-8-N-{6-methyl- 1H-pyrazolo[3,4- b]pyridin-3-yl}-5-N- [(2R)-1,1,1-trifluoropropan-2-yl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 44

(9S)-8-N-[4-(2-methyl- 1,3-thiazol-5- yl)pyridin-2-yl]-5-N- [(2R)-1,1,1-trifluoropropan-2-yl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 45

(9S)-8-N-{6-methyl- 1H-pyrazolo[3,4- b]pyridin-3-yl}-5-N-[(2S)-1,1,1-trifluoro-3- hydroxypropan-2-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide BA 46

(9S)-8-N-{6-methyl- 1H-pyrazolo[3,4- b]pyridin-3-yl}-5-N-[(2S)-1,1,1-trifluoro-3- hydroxypropan-2-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide AA 47

(9S)-3-methyl-8-N-{6- methyl-1H- pyrazolo[3,4-b]pyridin-3-yl}-5-N-[(2R)-1,1,1- trifluoropropan-2-yl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 5,8-dicarboxamide AA 48

(9S)-5-N-[(1R)-2,2- difluorocyclopropyl]-8- N-{6-methyl-1H-pyrazolo[3,4-b]pyridin- 3-yl}-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 49

(9S)-8-N-{5-[(2R)-2,3- dihydroxypropoxy] pyrazin-2-yl}-5-N-[(2R)-1,1,1-trifluoropropan-2- yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 50

(9S)-5-N-[(1R)-2,2- difluorocyclopropyl]-8- N-{4-[(2R)-2,3-dihydroxypropoxy] pyridin-2-yl}-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 51

(9S)-4-chloro-8-N-[4- (2-methyl-1,3-oxazol- 5-yl)pyridin-2-yl]-5-N-[(2R)-1,1,1- trifluoropropan-2-yl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 52

(9S)-5-N-[(1R)-2,2- difluorocyclopropyl]-8- N-{4-[(2R)-2,3-dihydroxypropoxy] pyridin-2-yl}-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 53

(9S)-5-N-[(1R)-2,2- difluorocyclopropyl]-8- N-{6-methyl-1H-pyrazolo[3,4-b]pyridin- 3-yl}-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 54

(9S)-8-N-{5-[(2S)-2,3- dihydroxypropoxy] pyridin-2-yl}-5-N-[(2R)-1,1,1-trifluoropropan- 2-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 55

(9S)-8-N-{6-[(2R)-2,3- dihydroxypropoxy] pyridazin-3-yl}-5-N-[(2R)-1,1,1-trifluoropropan-2- yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 56

(9S)-8-N-{5-[(2S)-2,3- dihydroxypropoxy] pyridin-2-yl}-3-methyl-5-N-[(2R)-1,1,1- trifluoropropan-2-yl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 57

(9S)-8-N-{4-[(2R)-2,3- dihydroxypropoxy] pyridin-2-yl}-3-methyl-5-N-[(2R)-1,1,1- trifluoropropan-2-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 5,8-dicarboxamide A A 58

(9S)-N-(5- fluoropyridin-2-yl)-5- [(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 4-carboxamide B 59

(9S)-N-(pyrazin-2-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 4-carboxamide B 60

(9S)-N-(pyridin-3-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 4-carboxamide B 61

(9S)-N-(pyridin-3-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 4-carboxamide B 62

(9S)-N-(pyrazin-2-yl)- 5-[(3S)-3- (trifluoromethyl)piperidin-1-yl]-1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 4-carboxamide B 63

(9S)-4-chloro-N- (pyridin-2-yl)-5-[3- (trifluoromethyl)phenyl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 64

(9S)-4-chloro-N-{6- [(2S)-2,3- dihydroxypropoxy] pyridin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 65

(9S)-4-chloro-N-{6- [(2R)-2,3- dihydroxypropoxy]pyridin- 2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 66

(9S)-4-chloro-5-(2- methylpyridin-4-yl)-N- (pyridin-3-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide B A 67

(9S)-4-chloro-N-{6- [(2S)-2,3- dihydroxypropoxy] pyrazin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 68

(9S)-4-methyl-5-(2- methylpyridin-4-yl)-N- (pyridin-3-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 69

(9S)-4-chloro-N-{2- [(2R)-2,3- dihydroxypropoxy] pyrimidin-5-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 70

(9S)-4-chloro-N-{6- [(2R)-2,3- dihydroxypropoxy] pyrimidin-4-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2,4,6-triene-8- carboxamide B A 71

(9S)-4-chloro-N- cyclopropyl-5-[3- (trifluoromethyl)phenyl]- 1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2(7),3,5-triene- 8-carboxamide A A 72

(9S)-4-chloro-N-{2- [(2S)-2,3- dihydroxypropoxy] pyrimidin-5-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 73

(9S)-4-chloro-N-{5- [(2R)-2,3- dihydroxypropoxy]pyrazin- 2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 74

(9S)-4-chloro-N-{5- [(2S)-2,3- dihydroxypropoxy]pyridin- 2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 75

(9S)-4-chloro-N-{4- [(2S)-2,3- dihydroxypropoxy] pyrimidin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 76

(9S)-4-chloro-N-{5- [(2R)-2,3- dihydroxypropoxy] pyridin-3-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 77

(9S)-4-chloro-N-{5- [(2S)-2,3- dihydroxypropoxy] pyrazin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 78

(9S)-4-chloro-N-{4- [(2R)-2,3- dihydroxypropoxy] pyrimidin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 79

(9S)-4-chloro-N-{6- [(2R)-2,3- dihydroxypropoxy] pyridin-3-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 80

(9S)-4-chloro-N-{5- [(2S)-2,3- dihydroxypropoxy] pyridin-3-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 81

(9S)-4-chloro-N-{2- [(2S)-2,3- dihydroxypropoxy] pyrimidin-4-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 82

(9S)-4-chloro-N-{4- [(2R)-2,3- dihydroxypropoxy]pyridin- 2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 83

(9S)-4-chloro-N-{2- [(2S)-2,3- dihydroxypropoxy]pyridin- 4-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 84

(9S)-4-chloro-N-{6- [(2S)-2,3- dihydroxypropoxy]pyridin- 3-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 85

(9S)-4-chloro-N-{2- [(2R)-2,3- dihydroxypropoxy]pyridin- 4-yl}-5-[3-(trifluoromethyl)phenyl]-, 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 86

(9S)-4-chloro-N-{6- [(2S)-2,3- dihydroxypropoxy] pyrimidin-4-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2,4,6-triene-8- carboxamide A A 87

(9S)-4-chloro-N-{5- [(2R)-2,3- dihydroxypropoxy]pyridin- 2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 88

(9S)-4-chloro-N-{2- [(2R)-2,3- dihydroxypropoxy] pyrimidin-4-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 89

(9S)-4-chloro-N-{5- [(2R)-2,3- dihydroxypropoxy] pyrimidin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2(7),3,5-triene- 8-carboxamide A A 90

(9S)-3-methyl-5-(2- methylpyridin-4-yl)-N- (pyridin-3-yl)-1,6,8-triazatricyclo[7.3.1.0²,⁷] trideca-2,4,6-triene-8- carboxamide A A 91

(9S)-N-{4-[(2R)-2,3- dihydroxypropoxy]pyridin- 2-yl}-4-methyl-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2,4,6-triene-8- carboxamide A A 92

(9S)-4-chloro-N-{5- [(2S)-2,3- dihydroxypropoxy] pyrimidin-2-yl}-5-[3-(trifluoromethyl)phenyl]- 1,6,8- triazatricyclo[7.3.1.0²,⁷]trideca-2,4,6-triene-8- carboxamide A A

Example 96

The present invention relates to Sirtuin Modulators, which are known inthe scientific literature for being useful for increasing lifespan of acell, and in treating and/or preventing a wide variety of diseases anddisorders, which include, but are not limited to, for example, diseasesor disorders related to aging or stress, diabetes, obesity,neurodegenerative diseases, cardiovascular disease, blood clottingdisorders, inflammation, cancer, and/or flushing as well as diseases ordisorders that would benefit from increased mitochondrial activity.

In addition to therapeutic potential, structural and biophysical studiesof SIRT1 activity and activation by small molecule sirtuin modualtorswould be useful in advancing understanding of the biological function ofsirtuins, mechanism of action of sirtuin activation and to aid indevelopment of assays that identify novel sirtuin modulators.

Based on the foregoing, the following literature references,respectively, are cited to demonstrate the utility of compounds of thepresent invention as Sirtuin Modulators and its interconnection withvarious diseases as exemplified or disclosed in the followingreferences:

-   1. Marcia C. Haigis and David A. Sinclair, Mammalian Sirtuins:    Biological Insights and Disease Relevance, Annu Rev Pathol. 2010; 5:    253-295.    -   Haigis and Sinclair teach:    -   “Aging is accompanied by a decline in the healthy function of        multiple organ systems, leading to increased incidence and        mortality from diseases such as type II diabetes mellitus,        neurodegenerative diseases, cancer, and cardiovascular disease.        Historically, researchers have focused on investigating        individual pathways in isolated organs as a strategy to identify        the root cause of a disease, with hopes of designing better        drugs. Studies of aging in yeast led to the of a family of        conserved enzymes known as the sirtuins, which affect multiple        pathways that increase the life span and the overall health of        organisms. Since the discovery of the first known mammalian        sirtuin, SIRT1, 10 years ago, there have been major advances in        our understanding of the enzymology of sirtuins, their        regulation, and their ability to broadly improve mammalian        physiology and health span. This review summarizes and discusses        the discovery advances of the past decade and the challenges        that will confront the field in the coming years (see, ABSTRACT,        therein and reference).”-   2. Gizem Donmezl et. al., SIRT1 and SIRT2: emerging targets in    neurodegeneration, EMBO Mol Med (2013) 5, 344-352.    -   Gizem Donmezl et. al., teaches:    -   “Sirtuins are NAD-dependent protein deacetylases known to have        protective effects against age-related diseases such as cancer,        diabetes, cardiovascular and neurodegenerative diseases. In        mammals, there are seven sirtuins (SIRT1-7), which display        diversity in subcellular localization and function. While SIRT1        has been extensively investigated due to its initial connection        with lifespan extension and involvement in calorie restriction,        important biological and therapeutic roles of other sirtuins        have only recently been recognized. Here, we review the        potential roles and effects of SIRT1 and SIRT2 in        neurodegenerative diseases. We discuss different functions and        targets of SIRT1 and SIRT2 in a variety of neurodegenerative        diseases including Alzheimer's disease (AD), Parkinson's disease        (PD) and Huntington's Disease (HD). We also cover the role of        SIRT1 in neuronal differentiation due to the possible        implications in neurodegenerative conditions, and conclude with        an outlook on the potential therapeutic value of SIRT1 and SIRT2        in these disorders (see, ABSTRACT, therein and reference).”-   3. Bracke et al., Targeted silencing of DEFB4 in a bioengineered    skin-humanized mouse model for psoriasis: development of siRNA    SECosome-based novel therapies; Exp Dermatol. 2014 Mar.; 23(3):    199-201. doi: 10.1111/exd.12321.    -   In particular, Bracke et al. teaches    -   “Psoriasis is a complex inflammatory skin disease that presents        a wide variety of clinical manifestations. Human β defensin-2        (hBD-2) is highly up-regulated in psoriatic lesions and has been        defined as a biomarker for disease activity. We explored the        potential benefits of targeting hBD-2 by topical application of        DEFB4-siRNA-containing SECosomes in a bioengineered        skin-humanized mouse model for psoriasis. A significant        improvement in the psoriatic phenotype was observed by        histological examination, with a normalization of the skin        architecture and a reduction in the number and size of blood        vessels in the dermal compartment. Treatment leads to the        recovery of transglutaminase activity, filaggrin expression and        stratum corneum appearance to the levels similar to those found        in normal regenerated human skin. The availability of a reliable        skin-humanized mouse model for psoriasis in conjunction with the        use of the SECosome technology may provide a valuable        preclinical tool for identifying potential therapeutic targets        for this disease.”-   4. Karline Guilloteau et al., Skin Inflammation Induced by the    Synergistic Action of IL-17A, IL-22 Recapitulates Some Features of    Psoriasis Oncostatin M, IL-1a, and TNF-a, J Immunol 2010;    184:5263-5270.-   Guilloteau et al. teaches:    -   “Keratinocytes play a crucial role in the regulation of skin        inflammation, responding to environmental and immune cells        stimuli. They produce soluble factors that can act in an        autocrine or paracrine manner on immune cells or directly on        aggressors. A screening of the activities of 36 cytokines on        keratinocyte gene expression identified IL-17A, IL-22,        oncostatin M, TNF-α, and IL-la as potent cytokines in inducing        cutaneous inflammation. These five proinflammatory cytokines        synergistically increased production of CXCL8 and b-defensin 2        (BD2). In addition, ex vivo studies on human skin explants        demonstrated upregulation of BD2, S100A7, and CXCL8 expression        in response to the same combination of cytokines. In vivo        intradermal injection of these five cytokines in mouse increased        CXCL1, CXCL2, CXCL3, S100A9, and BD3 expression, associated with        neutrophil infiltration. We confirmed and extended this        synergistic effect using quantitative real-time PCR analysis and        observed increased expression of nine chemokines and 12        antimicrobial peptides. Production of CXCL, CXCL5, and CXCL8 by        keratinocytes stimulated in the presence of this cytokine        combination was associated with increased neutrophil chemotactic        activity. Similarly, high production of BD2, BD3, and S100A7 was        associated with an increased antimicrobial activity. Finally,        the transcriptional profile observed in this in vitro model of        inflammatory keratinocytes correlated with the one of lesional        psoriatic skin. Our results demonstrate the important        potentiating activities of IL-17A, IL-22, oncostatin M, TNF-a,        and IL-1a on keratinocytes. This is particularly interesting in        the context of psoriasis where these cytokines are overexpressed        and could synergize to play an important role in upregulation of        chemokines and antimicrobial peptides production. The Journal of        Immunology, 2010, 184: 5263-5270 (see, ABSTRACT, therein and        reference)”.

Example 97 Description of Assays: PBMC Assay

Sirtuin 1 (Sirt1) is a homolog of silent information regulator 2 (Sir2)and a member of the NAD dependent class III histone deacetylase. Sirt1deacetylates lysine residues on histones, transcription factors andnonhistone proteins. Sirt1 has been shown to be involved in aging, cellcycle regulation, apoptosis, metabolic modulation and inflammation. Theactivation of Sirt1 causes deacetylation at lysine 310 of RelA/p65subunit of nuclear factor kB (NF-kB) transcriptional factor whichinhibits NF-kB transcription and down-regulates levels of TNFalpha.TNFalpha is a pleitotropic cytokine that is mainly produced bymacrophages and monocytes. TNFalpha is closely involved in immunedefense and chronic inflammation including Psoriasis. The expression oftype-1 cytokines such as TNFa was known to be increased in psoriaticskin and it plays important role in the etiology of psoriasis (Uyemura Ket al, 1993, J. Invest Dermatol, 101, p′701). Importantly, anti-TNFagent has been in clinical use for psoriasis. Therefore, Sirt1activators that induce a reduction in TNFa expression in inflammatorycells should have therapeutic effect in moderate to severe psoriaticpatients.

A PBMC/TNFalpha cell based assay was developed to identify activators ofSirt1 that inhibit the release of TNFalpha in response tolipopolysaccharide (LPS) stimulation of peripheral blood mononuclearcells (PBMC's). Briefly, PBMC's were stimulated by LPS, leading to anincrease in the production of TNFalpha secretion. TNFalpha protein levelwas measured by TNFalpha HTRF (homogeneous time resolved fluorescence)kit (CisBio, Inc). Cell lysis and TNFalpha detection were performedaccording to manufacturer's instructions. Sirt1 activators were testedin the presence of LPS to evaluate their inhibitory effect on TNFarelease and IC50 were determined in a dose-response experiment.

Beta-defensin 2 (bD2) ASSAY

Sirtuin is a family of NAD-dependent deacetylases which have broadphysiological functions and have been implicated in a number ofautoimmune and metabolic disorders including rheumatoid arthritis andtype I diabetes. Substrates of SIRT1 are diverse and includeinflammatory components with well established roles in innate andadaptive immune response such as NF□B, AP-1, FOXO, and p53.

Psoriasis is a chronic inflammatory skin disorder induced by genetic,autoimmune, and environmental factors. Lesions are characterized byhyperproliferation of keratinocytes in the epidermis and infiltration ofinflammatory cells resulting in chronic erythmatous plaques covered bywhite scales. Previous studies have shown that SIRT1 can impede theeffects of IL-22, a key cytokine in psoriasis, through direct inhibitionof STAT3 acetylation (Sestito et al, 2011). In addition, both SIRT1overexpression and resveratrol treatment (SIRT1 activation) can inducekeratinocyte differentiation (Blander et al, 2009).

Beta-defensin 2 (bD2) is an antimicrobial peptide that can be secretedfrom the epithelia where it acts as a chemoattractant for memoryT-cells, immature dendritic cells, and neutrophils. As such, bD2 is amajor part of the inflammatory response in the skin. Not only is bD2induced in lesional epidermal cells of psoriasis patients compared tonormal skin, but it is also a serum biomarker for disease severity inpsoriasis patients (Jansen et al, 2009; Kamsteeg et al 2009). Inaddition, bD2 may be genetically linked to psoriasis as a recent studyuncovered a significant association between increased beta-defensin genecopy number and psoriasis risk (Hollox et al, 2008). Of note, topicaldelivery of bD2 siRNA resulted in recovery of normal skin architectureand protein expression in a bioengineered skin-humanized mouse model forpsoriasis (Bracke et al, 2014).

An in vitro keratinocyte inflammation assay generated to mimic psoriaticinflammation was previously described (Guilloteau et al, 2010; Teng etal 2014). In these studies, a cytokine cocktail of IL-1alpha, IL-17A,IL-22, OSM, and TNFalpha (referred to as “M5”) was found to synergize toproduce a “psoriasiform” transcriptional profile in primary humankeratinocytes in vitro. In these studies, bD2 was one of the strongestresponders to the induction of keratinocyte inflammation.

Therefore, this assay was further developed in order to assess theefficacy of SIRT1 activator compounds for the topical psoriasis program.Specifically, conditions were optimized for an immortalized humankeratinocyte cell line (HaCaT) treated in vitro with the M5 cytokinecombination to induce psoriatic inflammation (as in reference above). Ina 48 hour time frame, bD2 secretion, as measured by a bD2 ELISA assay(Alpha Diagnostics), is significantly increased compared to unstimulatedkeratinocytes. This bD2 induction can be suppressed with treatment ofcompounds known to suppress psoriatic inflammation or, importantly, witha subset of SIRT1 activators. In parallel, cytotoxicity over the lengthof the 48 hour assay is ascertained by a CellTiter-Glo Luminescent CellViability Assay (Promega) to determine whether toxicity might play arole in bD2 response.

REFERENCES

-   Blander G, Bhimavarapu A, Mammone T, Maes D, Elliston K, Reich C,    Matsui M S, Guarente L, Loureiro J J. SIRT1 promotes differentiation    of normal human keratinocytes. J Invest Dermatol. 2009 January;    129(1): 41-9.-   Bracke S, Carretero M, Guerrero-Aspizua S, Desmet E, Illera N,    Navarro M, Lambert J, Del Rio M. Targeted silencing of DEFB4 in a    bioengineered skin-humanized mouse model for psoriasis: development    of siRNA SECosome-based novel therapies. Exp Dermatol. 2014 Mar.;    23(3): 199-201.-   Guilloteau K, Paris I, Pedretti N, Boniface K, Juchaux F, Huguier V,    Guillet G, Bernard F X, Lecron J C, Morel F. Skin Inflammation    Induced by the Synergistic Action of IL-17A, IL-22, Oncostatin M,    IL-1alpha, and TNFalpha Recapitulates Some Features of Psoriasis. J    Immunol. 2010 Mar. 24.-   Jansen P A, Rodijk-Olthuis D, Hollox E J, Kamsteeg M, Tjabringa G S,    de Jongh G J, van Vlijmen-Willems I M, Bergboer J G, van Rossum M M,    de Jong E M, den Heijer M, Evers A W, Bergers M, Armour J A, Zeeuwen    P L, Schalkwijk J. Beta-defensin-2 protein is a serum biomarker for    disease activity in psoriasis and reaches biologically relevant    concentrations in lesional skin. PLoS One. 2009; 4(3):e4725.-   Kamsteeg M, Jansen P A, van Vlijmen-Willems I M, van Erp P E,    Rodijk-Olthuis D, van der Valk P G, Feuth T, Zeeuwen P L,    Schalkwijk J. Molecular diagnostics of psoriasis, atopic dermatitis,    allergic contact dermatitis and irritant contact dermatitis. Br J    Dermatol. 2010 March; 162(3): 568-78.-   Sestito R, Madonna S, Scarponi C, Cianfarani F, Failla C M, Cavani    A, Girolomoni G, Albanesi C. STAT3-dependent effects of IL-22 in    human keratinocytes are counter regulated by sirtuin 1 through a    direct inhibition of STAT3 acetylation. FASEB J. 2011 Mar.; 25(3):    916-27.-   Teng X, Hu Z, Wei X, Wang Z, Guan T, Liu N, Liu X, Ye N, Deng G, Luo    C, Huang N, Sun C, Xu M, Zhou X, Deng H, Edwards C K 3rd, Chen X,    Wang X, Cui K, Wei Y, Li J. IL-37 ameliorates the inflammatory    process in psoriasis by suppressing proinflammatory cytokine    production. J Immunol. 2014 Feb. 15; 192(4): 1815-23.

Psoriasis & IL-17

Psoriasis is a chronic, relapsing, inflammatory autoimmune skin disorderwith a multi-factorial pathogenesis influenced by genetic,environmental, and immunopathologic factors (Griffiths C E et al.,Lancet 2007; 370:263-71). Psoriasis is characterized by recurrentepisodes of raised, well-demarcated erythematous oval plaques withadherent silvery scales. Histologically, the hallmark of psoriasis isthe presence of a thickened nucleated keratinocyte layer, withexaggeration of the rete pegs, caused by hyperproliferation ofkeratinocytes and dermal infiltration by activated T cells, neutrophils,and dendritic cells (Schon M P N. Engl. J. Med. 352: 1899-1912).

An accumulating body of evidence suggests psoriasis as a Th17-mediateddisease, driven by its signature cytokines IL-17 A, IL-17 F and IL-22.IL-22 induces proliferation of keratinocytes, whereas IL-17A stimulateskeratinocytes to secrete chemokines and other proinflammatory mediatorsthat recruit additional inflammatory cells, including neutrophils,dendritic cells, and innate lymphoid cells (Martin D A et al, J InvestDermatol 2013; 133:17-26).

The clinical validation of the IL-17 pathway in mediating psoriasis isdemonstrated by successful Ph3 studies that show significant improvementof disease using monoclonal antibody therapy targeting IL-17 (Langley etal., NEJM 2014). In addition, global transcription profiling inpsoriasis lesions following IL-17 inhibition suppressed multipleinflammatory factors from keratinocytes and leukocyte subsets to similarlevels as observed in non-lesional skin (Russell et al., J Immunol 2014,192: 3828-3836). Taken together, these findings support the role ofIL-17 in mediating psoriasis pathogenesis.

Method (Ex Vivo Skin Assay)

Stimulation of skin-resident immune cells in ex vivo human skin explantsusing a Th17 cytokine cocktail results in a dramatic upregulation ofTh17 related cytokines (IL-17A, IL-17F and IL-22), which establishesthis system as a human tissue-based model for psoriasis. The ability oftest compounds to modulate the expression of IL-17A, IL-17 F and IL-22was assessed using the ex vivo skin culture method post stimulation withTh17 cytokine cocktail.

Briefly, ex vivo human skin obtained from abdominoplasty surgery wasprocessed to remove fat and the tissue was dermatomed to −750 microns.Dermatomed skin was then cleaned in two serial rinses of 5-10 minuteseach in room temperature PBS containing an antibiotic/antimycoticsolution. The skin section was cut with disposable single-use biopsypunches to 10 mm diameter round sections, which were then placed in theupper chamber of a 0.4 μm PCF membrane transwell (Millicell #PIHP01250)containing 30 μl of a 64% bovine collagen solution (Organogenesis,#200-055) prepared with Cornification media. The skin samples wereallowed to set on the collagen solution for 30 min at 37° C. in ahumidified chamber. The skin samples on transwells were transferred to6-well plates (1 sample per well) and the lower chamber was filled with1 ml complete media (Cornification Media).

On the first day following abdominoplasty surgery, skin explants werecultured in Cornification media and allowed to incubate overnight at 37°C. Specifically, human skin explants (N=3 per condition) were stimulatedwith the Th17 cocktail (CD3, 1 μg/ml, CD28, 2 μg/ml, IL-1b, 10 ng/ml,IL-6, 5 ng/ml, TGFb, 1 ng/ml, IL-21, 10 ng/ml, anti-IL-4, 1 μg/ml andanti-INFg, 1 μg/ml). Test compound at 1,3 and 10 uM was added at thesame time as Th17 cocktail. Tissue was harvested 24 hrs after Th17activation and RNA was isolated for transcript quantification (IL-17A,IL-17F, IL-22) using qPCR.

Total RNA was isolated from ˜40 mg of tissue using Qiagen's Mini RNAIsolation kit (Cat #74106). Briefly, tissue was minced and homogenizedin the Precellys-24 machine using 300 μl of RLT buffer supplemented with1% 2-Beta-Mercapto-Ethanol at 6300 rpm for 30 seconds for 10 cycles witha 2-minute ice break. 490 μl of water containing 10 μl Proteinase K wasadded to the homogenate and digested at 55° C. for 15 minutes. Digestedtissue was spun down for 3 minutes at 10,000 G to pellet cell debri andthe supernatant was used for RNA isolation using Qiagen's RNeasy minicolumns according to manufacturer's protocol. Total RNA was quantifiedusing Nanodrop 2000 and analyzed on Agilent bioanalyser (filesattached). 1.4 μg of RNA was used as template in a 20 μl PCR volumeusing Invitrogen SuperScript VILO cDNA Synthesis kit (#11754-050) tocreate a cDNA template. Then cDNA was diluted 1:25 for the subsequentqPCR with the specific TaqMan probe for each gene to be quantified. RNAlevels of gene of interest's relative expression were calculated usingthe Delta Delta CT formula.

EQUIVALENTS

The present invention provides among other things sirtuin-modulatingcompounds and methods of use thereof. While specific embodiments of thesubject invention have been discussed, the above specification isillustrative and not restrictive. Many variations of the invention willbecome apparent to those skilled in the art upon review of thisspecification. The full scope of the invention should be determined byreference to the claims, along with their full scope of equivalents, andthe specification, along with such variations.

1.-18. (canceled)
 19. A compound or pharmaceutically acceptable saltthereof, selected from


20. A pharmaceutical composition comprising a compound of claim 19 and apharmaceutically acceptable carrier.
 21. The pharmaceutical compositionof claim 20 further comprising an additional active agent.
 22. A methodfor treating insulin resistance, a metabolic syndrome, metabolicdysfunctions, diabetes, or complications thereof, or for increasinginsulin sensitivity, comprising administering a compound or apharmaceutically acceptable salt thereof according to claim 19 to asubject in need thereof.
 23. A method for treating diseases or disordersresulting from diminished SIRT1 expression or activity, which comprisesadministering a compound or a pharmaceutically acceptable salt thereofaccording to claim 19 to a subject in need thereof.
 24. The methodaccording to claim 23 wherein the diseases or disorders resulting fromdiminished SIRT1 expression or activity are selected from aging orstress, diabetes, metabolic dysfunctions, neurodegenerative diseases,cardiovascular disease, cancer or inflammatory disease.
 25. The methodaccording to claim 23, wherein diseases or disorders are selected frompsoriasis, atopic dermatitis, acne, rosacea, warts, inflammatory boweldisease, Crohn's Disease, ulcerative colitis, osteoporosis, sepis,arthritis, COPD, systemic lupus erythematosus, phthalmic inflammation,alopetia, treatment of wounds, ocular disorders, dry eye, keratitis anduveitis.
 26. A method for treating insulin resistance, a metabolicsyndrome, metabolic dysfunctions, diabetes, or complications thereof, orfor increasing insulin sensitivity, comprising administering apharmaceutical composition according to claim 20 to a subject in needthereof.
 27. A method for treating diseases or disorders resulting fromdiminished SIRT1 expression or activity, which comprises administering apharmaceutical composition according to claim 20 to a subject in needthereof.
 28. The method according to claim 27 wherein the diseases ordisorders resulting from diminished SIRT1 expression or activity areselected from aging or stress, diabetes, metabolic dysfunctions,neurodegenerative diseases, cardiovascular disease, cancer orinflammatory disease.
 29. The method according to claim 27, whereindiseases or disorders are selected from psoriasis, atopic dermatitis,acne, rosacea, warts, inflammatory bowel disease, Crohn's Disease,ulcerative colitis, osteoporosis, sepsis, arthritis, COPD, systemiclupus erythematosus, phthalmic inflammation, alopetia, treatment ofwounds, ocular disorders, dry eye, keratitis and uveitis.